Early cognitive development relies on the sensory experiences that infants acquire as they explore their environment. Atypical experience in one sensory modality from birth may result in fundamental differences in general cognitive abilities. The primary aim of the current study was to compare visual habituation in infants with profound hearing loss, prior to receiving cochlear implants (CIs), and age-matched peers with typical hearing. Two complementary measures of cognitive function and attention maintenance were assessed: the length of time to habituate to a visual stimulus, and look-away rate during habituation. Findings revealed that deaf infants were slower to habituate to a visual stimulus and demonstrated a lower look-away rate than hearing infants. For deaf infants, habituation measures correlated with language outcomes on standardized assessments before cochlear implantation. These findings are consistent with prior evidence suggesting that habituation and look-away rates reflect efficiency of information processing and may suggest that deaf infants take longer to process visual stimuli relative to the hearing infants. Taken together, these findings are consistent with the hypothesis that hearing loss early in infancy influences aspects of general cognitive functioning. Infants learn about the world through their multimodal interactions with objects and other people in their environment –3. The aSeveral studies have shown that deaf children exhibit poorer performance in multiple nonverbal cognitive skills compared to their hearing peers. These include visual controlled attention , sequenDifferences between deaf and hearing individuals may initially arise from cortical cross-modal re-organization. In the absence of sound input, the auditory cortex responds to visual and somatosensory input ,15,16. VOne example, proposed by Smith and colleagues , is the An alternate explanation is that deaf children’s performance on cognitive and attentional tasks differs from that of hearing children because of their limited language experiences, rather than general cognitive abilities –22. EarlIn sum, a growing body of work provides evidence for differences in nonverbal cognitive abilities between deaf and hearing children, but the underlying sources remain unclear. Most of the research described above targeted preschool or school-age children, but no study has yet investigated the effects of hearing loss on cognitive abilities in young infants. As a result, little is known about when these differences between deaf and hearing children emerge.To address this gap, the current study aimed to compare visual habituation in infants with prelingual hearing loss and infants with typical hearing. Habituation is one of the earliest cognitive processes to emerge in development . VisualIf hearing loss is associated with differences in the early development of general cognitive abilities, we predicted that deaf infants may require more trials to reach habituation criteria, have slower habituation rates  and have lower look-away rates than hearing infants. To test these predictions, we compared the number of trials to reach the habituation criteria, growth slopes during habituation, and look-away rate between deaf and hearing infants using a visual habituation-oddity paradigm . For theM = 13.4, SD = 4.4). This range reflects the typical age range in which congenital deafness was identified in our population at the time of testing. For 19 of the infants, the etiology of hearing loss was unknown. The other four infants had a diagnosis of Mondini syndrome. Twenty-one of the infants used bilateral hearing aids. Sixteen infants came from families that used oral communication only and seven infants came from families that also used a signed language. Two infants had already received a CI: one was activated on the day of testing and one was activated one day prior to testing. Twenty-three additional deaf infants were tested  but excluded from analyses because of fussiness (n = 8), they fell asleep (n = 4), had known cognitive delays (n = 7), or did not meet the habituation criterion . This study was approved by the IRB committee of Indiana University.Deaf infants were diagnosed with bilateral severe-to-profound sensorineural hearing loss. All were scheduled to receive a CI or had a CI activated within 24 hours of participation in this study . All parn = 7) because it may indicate significant developmental, neurological, or cognitive delays in addition to hearing loss.Prior to participation in the study, all deaf infants underwent the typical evaluation for a CI at Riley Children’s Hospital in Indianapolis, Indiana. This included two standardized cognitive measures: The General Developmental Quotient from the Developmental Assessment of Young Children  and the M = 13.4, SD = 4.5) on the day of testing. Seventeen additional infants were tested  whose data were not included in the analyses because they did not complete testing (n = 15) or failed to habituate (n = 2). All hearing infants passed a newborn hearing screening, had no history of recurrent acute or chronic otitis media, and had no known developmental delays. Parents of all infants (deaf and hearing) provided informed written consent prior to inclusion in the study.Each infant in the deaf group was matched to a typically-developing hearing infant based on chronological age (+/- 1 month). The hearing group consisted of 23 infants  who ranged in age from 7.6 to 22.7 months  to measure their general visual attention over the course of the experiment. This is a method used in habituation studies to determine whether decreased looking times in infants are due to habituation to the stimuli or to general fatigue [A pre-test and post-test were conducted immediately before and after the experiment. Infants were presented with the geometric animation  rather than a fixed window of analysis  to maximize the opportunity for infants to reach the habituation criteria. Stimuli presentation continued until the infant reached the habituation criterion or up to a maximum of 18 trials .The dishabituation phase began immediately after the habituation criterion was reached. The experimenter was blind to when the experiment transitioned from habituation to the dishabituation phase. Like the habituation trials, each trial in this phase lasted for a minimum of one second and a maximum of 20 seconds. The dishabituation phase consisted of 14 trials: four novel trials and 10 familiar trials, which were presented in four continuous blocks. Following the procedure of , Block 1Throughout all phases of the experiment, looking time on each trial was calculated as the total time in seconds that infants were fixated on the visual stimulus. During the habituation phase, look-away rate was calculated by offline coding of each videotaped session by two independent coders who were blind to the infant’s hearing status. A look-away was defined as a gaze shift away from the visual stimuli during a trial that did not end that trial. Because trials only ended when infants looked away for more than one second (or until the 20s maximum) infants could make brief gaze shifts away from the screen (<1s) which did not end the trial. Look-away rate per minute was calculated as the total number of looks away from the stimulus during habituation, divided by the total looking time, and multiplied by 60s . Lastly,Habituation measures included the number of trials to reach criterion, growth slope over the first four trials, look-away rate, and dishabituation score. In the literature, the number of trials to reach habituation and dishabituation—i.e., novelty preference—has been shown to predict IQ later in development . Look-aReceptive and expressive language skills were assessed using the Auditory Comprehension and Expressive Communication subscales of the PLS-4, which were combined to yield a composite Total Language score. The PLS-4 measures sound awareness and vocalizations for infants younger than 12 months, and additionally includes items that measure word production and object knowledge for infants between 13 and 23 months of age. The Cognitive Scale of the Bayley Scales of Infant and Toddler Development  was usedTo examine relations between outcome measures and infant habituation behavior, we conducted Pearson bivariate correlations between the raw scores at each assessment year (pre- and post-CI) and habituation measures. Based on prior research, we hypothesized that longer time to habituate and lower look-away rates would reflect slower encoding of visual stimuli and would correspond to more protracted language and cognitive growth. We therefore predicted that time to habituate and look-away rate during habituation would correlate with scores on the Bayley Cognitive subscale at the pre-CI assessment, and that they would correlate with PLS-4 total language scores at pre- and/or post-CI assessments. We adopted a Bonferroni-adjusted alpha level of 0.01 (.05/4) in the evaluation of statistical significance. For thoroughness, we report the results of correlations between all habituation measures and outcome measures at both pre- and post-CI time points.ps > .34. Because our sample featured a large age range that extends beyond the typical range for habituation studies, we first examined whether results for the habituation measures differed for younger vs. older infants. To do so, we created a binary variable ‘Age’, to differentiate infants younger than 12 months vs. infants older than 12 months. Separate univariate analyses of variance (ANOVAs) were conducted for each habituation measure, with Hearing status (deaf vs. hearing) and Age (younger than 12 months vs. older than 12 months) as between-group factors. These analyses revealed no significant main effects or interactions with Age (ps > .18) for the number of habituation trials, mean habituation trial length, growth slopes, or dishabituation scores (ps > .15). Therefore, we collapsed across Age in subsequent analyses.We conducted a series of linear regression analyses to examine the effects of maternal education, unaided pure tone averages (PTA), and communication mode at time of test on habituation measures for the deaf infants. There were no effects of maternal education, communication mode, or unaided PTAs on any variable, ps > .25), although some infants did show decreased looking during the post-test compared to the pre-test (n = 8 in the deaf group and n = 8 in the hearing group). There were also no differences between groups on the post-test, p = .46. However, deaf infants showed significantly longer looking times during the pre-test than hearing infants, F = 4.91, p = .03, d = .66. See There were no differences in overall looking time between pre- and post- test looking times within each infant group separately  = 4.40, p = .04, d = .63  = 4.22, p < .05, d = .65  = 0.59, p = .44.Deaf infants required a greater number of trials to reach the habituation criterion,  d = .63 , and dem d = .65 . Total l studies . The gro p = .02 . Togethet(22) = 3.99, p < .01, d = 1.18, hearing t(22) = 2.49, p = .02; d = .73. Thus, although the deaf infants took longer to habituate to the stimuli, there were no group differences in subsequent responsiveness to a novel visual stimulus.During the dishabituation phase, both groups of infants demonstrated a significant novelty preference, looking longer to the novel stimulus relative to the familiar stimulus across trials: deaf F = 4.75, p = .04, d = .65. Together, these findings reveal that deaf infants are slower to habituate to a visual stimulus and show lower look-away rates during habituation than hearing infants, suggesting that hearing loss may be associated with differences in basic cognitive processing of visual stimuli during infancy.Differences in looking times extended throughout the dishabituation phase. Deaf infants continued to look significantly longer than the hearing infants did across the novel and familiar trials, r = -.45, p = .011, n = 20. Deaf infants who habituated faster to a visual stimulus displayed higher language scores  were excluded from the current study. For the deaf infants, we also found that time to habituate correlated with language scores at the time of testing, indicating that infants who habituated more slowly had lower language levels. Together, these findings reveal differences in general processing of non-auditory visual information between deaf and hearing infants, at a younger age than has previously been tested. They also illustrate that prelingual hearing loss results in consequences that extend beyond the auditory system.Our findings suggest that deaf infants’ visual processing is slower or less efficient than that of hearing infants. Habituation is thought to reflect memory and the speed at which infants encode a stimulus, which are predictive of later cognitive and language skills . Look-awAnother possible interpretation of our results is that longer looking times and lower look-away rates reflect differences in sustained attention, rather than efficiency of information processing. In Quittner et a. (2012), deaf children with CIs showed faster improvement in visual attention skills—specifically, perceptual sensitivity and attentional control—over the course of a year relative to deaf children who used hearing aids . The autThe ability to sustain visual attention could be an advantage for the deaf infants rather than a deficit, as they may be acquiring richer representations of the visual stimuli than their hearing peers may. However, our data show that infants with fewer trials to habituate scored higher on standardized language assessments, suggesting that longer habituation times do not correspond to better spoken communication abilities. Given the established links between the development of attention and oral communication skills, our findings do not support the hypothesis that slower habituation reflects enhanced attention to the stimuli. Moreover, a review of the past 30 years of research on the visual abilities of deaf individuals suggests that their superior performance on visual tasks is best characterized by enhanced reactivity to visual events, rather than enhanced perceptual representations . HoweverOur findings are also consistent with prior evidence that habituation length and look-away rates during habituation are complimentary measures of processing efficiency . ResearcOne explanation for slower visual processing in the deaf infants is their limited auditory experiences. Limited exposure to auditory input from birth has been proposed to result in delays in general cognitive development . AccordiAn alternative explanation is that deaf infants’ cognitive development may be protracted because of delayed language development. Language and cognitive skills develop interdependently and likely share common underlying mechanisms in both deaf and hearing children ,49. ConsIt remains difficult to untangle whether differences in visual processing in deaf infants and children result from delays in cognitive function, language exposure, or both. In prior research, one potential confound is that some of the tasks used to examine cognitive performance in deaf and hearing individuals may rely on verbal rehearsal strategies, therefore creating a natural disadvantage for the deaf group . The curTo isolate the contribution of auditory experience to visual attention, independent of language, Dye & Hauser  conducteOne additional possibility to consider is that structural-functional changes in the brain could underlie differences in processing speed or efficiency. As described in the introduction, deprivation in one sensory modality results in cross-modal plasticity throughout the brain. In the case of hearing loss, there is evidence to suggest that these cross-modal changes—in which the auditory cortex assumes some of the functions of visual cortices—can result in both adaptive gains but also maladaptive outcomes . For insOne limitation of the current study is the relatively wide age range of our sample, which is a common characteristic of conducting research with deaf infants prior to cochlear implantation. Because of this wide age range, the mechanisms driving visual habituation behaviors could potentially shift. However, our aim was to examine the difference in behavioral responses to unimodal visual stimuli between deaf and hearing infants, rather than to make claims about the potential physiological mechanisms underlying visual habituation. Future work should replicate these findings with a narrower age range.This study reveals differences in visual habituation, a building block of cognitive development, between deaf and hearing infants. Deaf infants required a greater number of trials to habituate, showed slower rates of habituation, and had lower look-away rates during habituation than age-matched hearing infants. These data provide converging evidence for slower or less efficient processing of the visual stimuli than their hearing counterparts. In addition, these measures of visual habituation correlated with the deaf infants’ performance on standardized tests of language skills. Together, our findings provide the first evidence to suggest that differences in processing visual stimuli between deaf and hearing children emerge early in infancy, prior to advanced language acquisition. Our findings also support the notion that early differences in visual information processing may explain some of the variability in deaf children’s neurocognitive development. Given the predictive relationship between habituation behaviors in infancy and later cognitive development ,47, it iS1 File(SAV)Click here for additional data file.


Fabry Disease (FD) is an X-linked hereditary lysosomal storage disorder which leads to a multisystemic intralysosomal accumulation of globotriaosylceramid (Gb3). Besides prominent renal and cardiac organ involvement, patients commonly complain about vestibulocochlear symptoms like high-frequency hearing loss, tinnitus and vertigo. However, comprehensive data especially on vertigo remain scarce. The aim of this study was to examine the prevalence and characteristics of vertigo and hearing loss in patients with FD, depending on renal and cardiac parameters and get hints about the site and the pattern of the lesions.Single-center study with 57 FD patients. Every patient underwent an oto-rhino-laryngological examination as well as videonystagmography and vestibular evoked myogenic potentials (VEMPs) and audiological measurements using pure tone audiometry and auditory brainstem response audiometry (ABR). Renal function was measured by eGFR, cardiac impairment was graduated by NYHA class.More than one out of three patients (35.1%) complained about hearing loss, 54.4% about vertigo and 28.1% about both symptom. In 74% a sensorineural hearing loss of at least 25 dB was found, ABR could exclude any retrocochlear lesion. Caloric testing showed abnormal values in 71.9%, VEMPs were pathological in 68%. A correlation between the side or the shape of hearing loss and pathological vestibular testing could not be revealed.Hearing loss and vertigo show a high prevalence in FD. While hearing loss seems due to a cochlear lesion, peripheral vestibular as well as central nervous pathologies cause vertigo. Thus, both the site of lesion and the pathophysiological patterns seem to differ.The online version of this article (10.1186/s13023-018-0882-7) contains supplementary material, which is available to authorized users. Fabry disease (FD) is an X-linked lysosomal storage disorder which is characterized by a reduced or absent enzyme activity of α-galactosidase A. This leads to an intralysosomal accumulation of globotriaosylceramid (Gb3), which results in tissue damage of kidneys, heart and the nervous system –4. The iSince the introduction of enzyme replacement therapy (ERT) in 2001, a reduction of Gb3 storage in kidneys and the heart could be shown –22. FurtDespite the profound impact of hearing loss and vertigo on patients’ individually experienced health-related quality of life, comprehensive data supporting the development of new guidelines for the monitoring and treatment of Fabry disease remain scarce due to prognostic domination of other organs. The aim of this current study was to evaluate the prevalence and characterize the patterns of vertigo and hearing loss in dependence of other Fabry-typical organ manifestations and to get hints about the site and the pattern of the lesions.Fifty-seven FD patients , who attended the Department of Oto-Rhino-Laryngology, Plastic, Aesthetic and Reconstructive Head and Neck Surgery in Würzburg, were investigated between 04/2012 and 11/2016. Informed oral and written consent had been obtained appropriate to the decision of the institutional review board of the medical department Würzburg . All patients were recruited from the Würzburg Fabry Center for Interdisciplinary Therapy (FAZIT) within the scope of routine check-ups irrespective of any ENT symptoms or comorbidities. Inclusion criteria were age ≥ 18 years and confirmed diagnosis of FD by DNA testing and α-galactosidase A assay.Medical history was taken of all patients before a complete oto-rhino-laryngological examination. Especially, they were asked about hearing loss, tinnitus and vertigo as well as ototoxic medication, noise exposure or infections.2 [Glomerular filtration rate (CKD-EPI equation) was measured to estimate renal function with following graduation: ≥90, 60–89, 30–59 and ≤ 29 ml/min/1.73 m2 . Cardiac2 . Patient2 .Audiological measurements were performed with calibrated instruments in a sound-proofed room (DIN EN ISO 8253). The audiological evaluation included standard pure-tone audiometry , conducted with a clinical audiometer in 5-dB steps. Hearing thresholds were then averaged in 4-pure tone average  and a modified 6-pure tone average , summarizing all values and dividing by 4 resp. 6, so every threshold carries equal weight. Values 10 dB above normative hearing thresholds were considered abnormal .Otoacoustic emissions  were performed in each patient. Furthermore, auditory brainstem response audiometry (ABR) was performed using Eclipse - ASSR EP15/EP25  in 56/57 patients. Click stimuli were presented at intensities between 10 and 100 dB HL and responses were then averaged and the ABR threshold was visually determined where wave V showed the smallest response amplitude.Videonystagmography (VNG) with recording of spontaneous nystagmus (SPN) and caloric testing with warm (44 °C) and cold (30 °C) water or air  were performed. Results were considered abnormal when canal paresis factor (CP) was above 25%. Cervical Vestibular Evoked Myogenic Potentials (cVEMPs) were recorded ipsilaterally from the tonically activated sternocleidomastoid muscle by surface electrodes. Ocular VEMPs (oVEMPs) were detected contralaterally by surface electrodes inferior the eye, while the patient was looking upward. Stimulation was carried out with monaural clicks of 100 dB and a rate of 5,1 Hz each via insert tips. Each measurement was performed twice and results averaged.p < 0.05).A normal distribution was not found using Shapiro-Wilk test, so Kruskal-Wallis test and pairwise Wilcoxon’s rank sum test were applied. Statistical significance was set at the 95% confidence level and above  at the time of examination. Of these 20 patients, 18 (90%) complained about one or more episodes of sudden hearing loss which was asymmetric, and 2 patients reported slowly progressive hearing loss. Another 5 patients described hearing loss in the past, which had subjectively fully recovered. Tinnitus was described by 43.9% . Vertigo was reported by 54.4%, whereof 17 patients had intermittent, 2 had permanent and 12 had vertigo which could be triggered. In 15.8% vertigo was described as rotary and in each 19.3% it was named staggering or non-directional  with a mean period of time on medication of 5.97 years (range 1–15 years).2. According to the NYHA score, 22 subjects were scored within class 0, 13 patients in class 1, 16 patients in class 2 and 6 patients in class 3. 38 patients were assigned to the ‘classical’ FD mutation group and 7 patients had a ‘late onset’ mutation like N215S; 12 patients had mutations that – based on current knowledge – could not be specified.Renal function was determined with the eGFR grouped according to KDIGO categories. Twenty-one subjects showed a value of ≥90, 20 patients a value of 60–89, 13 patients a value between 30 and 59 and 3 male patients a value of ≤29 ml/min/1.73 mPure-tone audiometry showed an asymmetric high-frequency sensorineural hearing loss. Conductive or combined hearing loss did not occur. In 42 patients (73.7%) we found a sensorineural hearing loss ≥25 dB HL in at least one frequency while only 18 (42.9%) of these patients subjectively had symptoms of hearing loss. Men were affected more severely than women. In two patients who reported hearing disability we found normal values in pure-tone audiometry < 25 dB HL in all frequencies.4-PTA of the better ear according to the WHO classification of disability due to hearing loss showed pathological results (> 25 dB) in 6 patients . Inspecting the bad ear by reason of a markedly asymmetric hearing loss, 17 patients (29.8%) showed at least a slight impairment . In higher frequencies above 2 kHz, the degree of hearing loss was significantly depending on the severity of renal and cardiac function . The categorization of the patients to the groups ‘classical’ versus ‘late onset’ mutations also did not reveal any difference.Serum lyso-Gb3 levels had no influence on 4-PTA values , a statistically significant increase of ABR-thresholds could be demonstrated between subgroups . Of these, 26 reported vertigo and 24 were free of symptoms. CVEMPs weren’t derivable in 5 ears. Mean latencies of p1/p13 were at 12.0 ms and of n1/n23 at 21.2 ms. Peak-to-peak amplitudes (n1-p1) were 45.5 μV. Pathological cVEMPs values were found in 24 individuals, which can indicate a malfunction of sacculus respectively the inferior vestibular neve: a reduction of amplitude was seen in 17 patients, an extension of latency in 12 patients (partly with overlaps).OVEMPs could not be elicited in 10 patients (20%), which was not rated as pathological as in previous studies a lack of oVEMPs in up to 50% of healthy individuals was already described . UnilateWith increasing age , a prolongation of latencies and decrease of amplitudes in cVEMPs and oVEMPs were detected Table .Table 2L-d showing only cVEMPs).Depending on the severity of renal failure (measured by GFR) respectively cardiac insufficiency (rated on NYHA class), a partly significant decrease of cVEMP and oVEMP amplitudes (peak-to-peak n1-p1) was found. A prolongation of p1 latency (cVEMPs) and n1 latency (oVEMPs) could tendentially be depicted, not being significant . There likewise was no difference seen between patients with ‘classical’ and ‘late onset’ mutations.Lyso-Gb3 levels also had no influence on cVEMP and oVEMP values . There was also an increase in latencies . Regarding oVEMPs, a decrease in amplitude and an increase in latencies . The results suggest that patients who suffer from sensorineural hearing loss are more likely to have vestibular lesions. In patients with vertigo, smaller amplitudes  as well as higher latency values  were registered than in those without vertigo.Modified 6-PTA as parameter of hearing loss revealed a significant negative correlation with the amplitude of cVEMPs in linear regression analysis Fig. . A loweries Fig. , which wies Fig. /d was alIn this group of 57 FD patients, a high incidence of sensorineural high-frequency hearing loss (73.7%) was apparent. Previously published studies, mostly small case studies, reported an incidence ranging between 19 and 87% , 23, 29,The severity of hearing loss is significantly correlated to the function and injury of both kidney and the heart . One limSince retrocochlear pathology could be excluded by ABR, it is supposed that the lesion is located in the inner ear , 16. ThiVertigo seems to follow a more complex pattern. The incidence of vertigo was 54.4% and could occur separately or in combination with hearing loss or tinnitus. FD patients showed a higher incidence compared to a telephone survey with more than 8000 participants conducted in Germany in 2003, where 22.4% of men and 36.2% of women reported having suffered at least once from dizziness . Data abIn 71.9% pathological nystagmus reactions were recorded in VNG and CP was increased > 25% in 29.8%. Most common findings were a one-sided reduction, which suggests a peripheral vestibular lesion, or a bilateral inhibitory deficit, which is more an expression of a central genesis. Other authors showed abnormal VNG results between 17.5%  and 21% CVEMPs were pathological in 48% and oVEMPs in 44%. Amplitude reduction as sign of peripheral vestibular or otolithic origin was observed 17 times in cVEMPs and 12 times in oVEMPs. An extension of latency as sign of a neurogenic or central pathology was found 12 times in cVEMPs and 13 times in oVEMPs. These results are comparable to a recently published study with a mixed-gender group of 36 Fabry patients . For theIncreasing renal or cardiac dysfunction did not lead to significant changes in VNG. In VEMPs, a tendency towards amplitude reduction or latency extension was observed, although only partially significant. However, individual groups (GFR / NYHA) in some cases had very different distributions. In future work, further data collection for more representative groups is important.Although serum lyso-Gb3 has been revealed to be an important biomarker for staging FD , lyso-GbSeveral studies have shown that the type of mutation substantially affects organ involvement , 38, 39.Of course, also other common and non Fabry-related reasons of dizziness like cardiac insufficiency  need to VEMPs should always be interpreted in the overall context and, if possible, combined with other vestibular tests like VNG or video head impulse testing. Since values in individuals vary relatively widely and depend on the type of stimulus  and the intensity of stimulation, narrowly set standard values are difficult to ascertain –44. HoweAt the time of presentation, 35.1% of all FD patients perceived subjective hearing loss, 56.9% reported dizziness symptoms and 28.1% reported a combination of both symptoms. However, it was remarkable that a simultaneous onset of symptoms could rarely be indicated. In addition, 33.3% of all patients noticed hearing impairment in the sense of an acute hearing loss, whereas only 2.9% reported a sudden onset of dizziness, as in the case of a vestibular neuritis. On the other hand, in cases of common presence, there was no correlation between the side of a measurable hearing loss and the side of a pathological caloric or electromyographic measurement. This raises the question whether these are different types of lesions or different pathophysiological causes . Other authors reached similar results , 36, notIn contrast to the audiological results, a clear pathophysiological pattern could not be identified in the diagnosis of vertigo, so that combined peripheral and central vestibular pathologies have to be assumed. Further clinical and pathohistological studies are necessary to decipher the pathophysiology of vestibulocochlear symptoms in Fabry’s disease.High-frequency hearing loss and vertigo are common in FD patients. Hearing loss is due to a cochlear lesion without any signs of retrocochlear pathology. Vertigo seems to be caused by peripheral vestibular as well as central nervous pathologies. The site of lesion and the pathophysiological pattern seem to differ. Every FD patient should obtain an extensive audiological and vestibular testing regularly.Additional file 1:Pure Tone Audiometry in relation to renal and cardiac function. Pure Tone Audiometry (AC) of the bad ear shows that hearing loss above 2 kHz is depending on the grade of GFR and NYHA. (PDF 1355 kb)Additional file 2:ABR thresholds vs. GFR and NYHA. According (A) to progredient renal dysfunction  and (B) cardiac dysfunction , a statistically significant increase of ABR thresholds could be demonstrated between single groups. (PDF 1390 kb)


Hearing loss is a common sensory disorder that has been a serious concern globally. It is recently estimated that around 466 million people worldwide have disabling hearing loss, including 34 million children, and this number will increase to over 900 million by 2050 (according to WHO report 2018). The majority of hearing disorders occur due to the death of either inner ear hair cells (HCs) or spiral ganglion neurons (SGNs), thus leading to sensorial neural hearing loss (SNHL). SNHL is known as the most common form of hearing disorder that comprises about 85% of all hearing loss cases. This type of damage is induced by a variety of reasons such as inner ear trauma, ischemia, ototoxic drugs, noise exposure, inflammation, viral infections, genetic deficits, autoimmunologic reaction, and aging. SNHL is generally not reversible due to the lack of regeneration capacity of HCs and SGNs. However, various recent studies have determined that the HCs and SGNs hold a regenerative potential and it is possible to find the cure for SNHL in the near future. This is supported by the clear understanding of the genetical control and signaling pathways involved in the development of HCs and SGNs and their functions, the regenerative potential of residing adult stem cells, and the development of gene therapy and the clinical trials of new pharmaceutical compounds on damaged cochleae. Last year, we have published the first special issue of “Hearing Loss: Reestablish the Neural Plasticity in Regenerated Spiral Ganglion Neurons and Sensory Hair Cells”, and this year in this second special issue, we are presenting a new series of articles to report the most recent advances in several major areas as summarized below: HC development, HC damage and protection, HC regeneration, SGN development and protection, inherited hearing loss, and inner ear drug delivery.In Vivo”) for the first time experimentally demonstrate the backward traveling wave theory by measuring the phase spectra of the basilar membrane vibration at multiple longitudinal locations along the cochlea. X. Cheng et al. (“Modulation of Glucose Takeup by Glucose Transport on the Isolated OHCs”) report that glucose is transported into OHCs via glucose transporter 1 and 4, which are mainly expressed on the lateral wall of OHCs and the glucose antagonist and ATP regulate this energy transport mechanism. S. Liu et al.  determine that Pax2, Sox2, and Prox1 have differential and overlapping temporospatial expression patterns during the development of vestibular and auditory sensory organs in mice.A. Chang et al.  report that the early environmental sounds promote functional maturation of HCs. Acoustic environment significantly decreases the ABR threshold, increases prestin expression in outer HCs, and enhance maturation of ribbon synapses in the postnatal mouse cochleae. P. Chen et al.  demonstrate that the microglial-like cells are present in the developing mouse cochlea, and these cells go through the drastic morphological and distributional changes during the postnatal cochlear development. Also, these cells might participate in the maturation and remodeling of the cochlea. F. Chen et al.  explore the pathogenesis of superior semicircular canal dehiscence (SSCD) in the guinea pig model and report that the bony fenestration of the superior semicircular canal mimics the hearing loss pattern of SSCD patients. J. Hong et al.  investigate that the N-methyl-D-aspartate receptors regulate the number and distribution of inner HC ribbon synapses after gentamicin-induced ototoxicity and their inhibition by antagonist minimized the drug-induced ototoxicity, and thus maintain the integrity of ribbon synapses. L. Xia et al. (“Comparison of Acceptable Noise Level Generated Using Different Transducers and Response Modes”) compare the acceptable noise level (ANL) in 20 mandarin subjects with normal hearing. The author obtained ANL through different methods and determined that the ANL in normal hearing listeners may not be affected by different modes of presentation. M. Waqas et al.  provide a brief review about the mechanisms involved in the HC loss after noise-induced trauma and discuss the recent HC protection strategies to prevent and recover hearing function in mammals after noise-induced damage. X. Cheng et al.  report that the combination of electric and acoustic hearing significantly improves the perception of music and Mandarin tones in pediatric cochlear implant patients. X. Ding et al.  determine the association between tinnitus and sudden sensorineural hearing loss (SSNHL). The authors found that tinnitus can be ameliorated by the successful treatment of SSNHL. G. Li et al.  provide a brief review on the autoimmune mechanisms involved in SSHL and discuss the role of immunosuppressive drugs in immune therapy. N. Zhao et al.  report that the age-related hearing loss causes an increase in PnC sensitivity that in turn enhances acoustic startle responses in C57 mice. B. Li et al.  investigate the extent of SNHL at high frequencies influences the ability to recognize compressed speech of lower frequencies in hearing loss patients.M. Tang et al.  provide a comprehensive review to address the current challenges and problems in stem cell transplant-based treatments in the inner ear against deafness and present a critical viewpoint about electrical stimulations as a physical factor to modulate stem cell behavior and promote stem cell therapy to treat hearing loss.J. Li et al.  determine that the type II SGNs participate in the contralateral suppression of the medial olivocochlear reflex after selectively inducing apoptosis in the type I SGNs using ouabain treatment.MYO15A Mutations Identified in One Chinese Family with Autosomal Recessive Nonsyndromic Hearing Loss”) report three MYO15A variants c.3971C>A (p.A1324D), c.4011insA (p.Q1337Qfs∗22), and c.9690+1G>A. These variants are absent in 200 normal controls and cosegregated with hearing disability in this family. X. Wang et al. (“A Novel p.G141R Mutation in ILDR1 Leads to Recessive Nonsyndromic Deafness DFNB42 in Two Chinese Han Families”) identify a novel p.G141R homozygous mutation in ILDR1 gene that may be the genetic cause of deafness in two unrelated Chinese Han families. X. Wu et al.  report that the novel compound heterozygous missense mutation c.4472C>T p.T1491M and c.1973T>C p.V658A in PTPRQ gene is the genetic cause of recessively inherited sensorineural hearing loss in a Chinese family.H. Du et al. (“Identification of Binding Partners of Deafness-Related Protein PDZD7”) determine the eleven novel PDZD7-binding proteins through yeast two-hybrid screening that are expressed in the inner ear. Most of the new PDZD7-binding partners such as TRIM35, CADM1, AMOT, BLZF1, Numb, KCDT10, CCDC27, and TRIP11 have not been reported before and will help to understand the role of PDZD7 in hearing transduction. P. Li et al. (“Knock-In Mice with Myo3a Y137C Mutation Displayed Progressive Hearing Loss and Hair Cell Degeneration in the Inner Ear”) report that Myo3a kinase domain Y137C mutant mice have an elevated hearing threshold, degenerated inner ear HCs, and structural abnormality in HCs stereocilia after 6 months of age, thus Myo3a is essential for maintaining the intact structure of HC and normal hearing function. S. Hu et al. (“Genetic Etiology Study of Ten Chinese Families with Nonsyndromic Hearing Loss”) identify novel pathogenic variants in six Chinese families with a hereditary hearing loss by targeted next generation sequencing. F. Zhang et al.  synthesize a new nanoparticle capsule that can be used as a drug delivery route for the gentamicin transfer at the specific site in the inner ear. The authors also determine the sustained release capacity of gentamicin from the capsule by We believe that the studies included in this second special issue of “Hearing Loss: Reestablish the Neural Plasticity in Regenerated Spiral Ganglion Neurons and Sensory Hair Cells” provide important insights into cochlear physiology and pathology as well as the important progress in technology that can be translated into clinical application of the medical treatment of cochlear damage in SNHL. We wish that this special issue will represent a significant contribution in the effort to achieve effective protection and treatment of hearing loss in the near future.


Optimal therapy for colorectal carcinoma (CRC), a frequently diagnosed malignancy, does not exist. Some of colicins and microcins, ribosomally synthesized peptides by gramnegative bacteria, have shown significant biological activity specifically against different cancer cells in vitro and in vivo conditions. The aim of this prospective study was to evaluate natural colicin and microcin production by large intestinal mucosal bacteria in each stage of colorectal neoplasia and in those with a history of colorectal neoplasia.A total of 21 patients with non-advanced adenoma , 20 patients with advanced colorectal adenoma , 22 individuals with CRC (9/22 with current and 13/22 with history of CRC) and 20 controls were enrolled. Mucosal biopsies from the caecum, transverse colon and the rectum were taken during colonoscopy in each individual. Microbiological culture followed. Production of colicins and microcins was evaluated by PCR methods.A total of 239 mucosal biopsies were taken. Production of colicins and microcins was significantly more frequent in individuals with non-a-A, a-A and CRC compared to controls. No significant difference in colicin and microcin production was found between patients with current and previous non-a-A, a-A and CRC. Significantly more frequent production of colicins was observed in men compared to women at the stage of colorectal carcinoma. A later onset of increased production of microcins during the adenoma-carcinoma sequence has been observed in males compared to females.Strains isolated from large intestinal mucosa in patients with colorectal neoplasia produce colicins and microcins more frequently compared to controls. Bacteriocin production does not differ between patients with current and previous colorectal neoplasia. Fundamental differences in bacteriocin production have been confirmed between males and females. Colorectal carcinoma (CRC) is after the lung and prostate cancer the third most frequently diagnosed malignancy in males and the second most common malignancy after the breast cancer in females. There were 1006.000 new CRC diagnoses in males and 794.900 in females in 2018 worldwide. A total of 474,600 CRC related deaths in males and 387,100 in females were reported in 2018 worldwide [The conventional strategies used for treatment of CRC are endoscopy (reserved for early carcinoma up to stage T1sm1), chemotherapy, radiotherapy and surgery. The major disadvantage of chemotherapy, which is the main choice of treatment, is the non-specific mode of action associated with significant toxicity and frequently observed secondary resistance of cancer cells towards chemotherapeutic agents , 3.14 microbes and to about 1000 bacterial species [The human large intestine is a home to approximately 10 species . The con species , neverth species , 7, 8.Bacteriocins are substances ribosomally synthesized by bacteria. They exhibit antimicrobial activity towards other bacterial species –11. MajoSome of colicins and microcins, bacteriocins produced by gramnegative bacteria, have shown biological activity against different cancer cells in vitro and in vivo conditions , 14. TheThe aim of this prospective study was to evaluate natural colicin and microcin production by large intestinal mucosal bacteria in each stage of colorectal neoplasia and in those with a history of colorectal neoplasia.A total of 63 patients with colorectal neoplasia and 20 controls were enrolled into the study. Three groups of patients with colorectal neoplasia were included: individuals with non-advanced colorectal adenoma, non-a-A , patients with advanced colorectal adenoma, a-A  and individuals with CRC . Control group consisted from 7 men and 13 women (mean age 57 ± 14). Patients with colorectal neoplasia were in average risk of developing a sporadic colorectal carcinoma. Advanced colorectal adenoma is defined as an adenoma with low grade dysplasia larger than 10 mm and/or containing high grade dysplasia being of any size and/or adenoma of any size with villous component [Among controls, individuals with average risk for colorectal carcinoma, with normal findings on colonoscopy and with negative history of colorectal neoplasia and/or inflammatory bowel disease were included.Polyethylene glycol was used most frequently for bowel preparation. A total of 40% controls, 62% of patients with a non-advanced adenoma, 50% of individuals with an advanced adenoma and 64% of patients with a colorectal carcinoma were prepared with polyethylene glycol. The remaining patients were prepared with sodium phosphate or sodium picosulfate or sulphate salts. was performed. Obtained bacterial strains from the Enterobacteriaceae family were frozen subsequently.We used the original methodology reported by our group . MucosalE. coli K12-ROW, E. coli C6 (phi) and Shigella sonnei 17 were used for the assessment of bacteriocin production. Bacteriocin production was also confirmed by PCR methods.Production of colicins and microcins was evaluated and investigation of bacteriocin type was carried out: frozen bacterial strains were plated on Petri plates with either 1.2% Trypton Yeast agar or Nutrient agar. After a 48-h culture plates were treated with chloroform vapour for 30 min. Trypton Yeast agar with an indicator strain was added. Indicator strains of Determination of type of bacteriocin was accomplished by PCR methods using specific primers for the detection of colicin and microcin genes . AltogetObtained data were tested statistically by means of descriptive statistics and Fisher’s exact test with STATISTICA software, version 13, 2013, Tulsa, OK, USA.All patients enrolled into the study were adequately informed, supplied and signed an informed consent. The project was approved by the Joint Ethical Committee  under the number 201107 S54. For all data obtained, all personal identification information was removed in compliance with the Czech laws for protection of confidentiality.Twenty controls, 21 patients with non-a-A, 20 individuals with a-A and 22 patients with CRC were included. A total of 239 mucosal biopsies were taken ..Colicin producing strains were identified in 19% (10/52) controls, 49% (31/63) non-a-A, 43% (26/60) a-A and in 61% (39/64) CRC; Fig. 1.Microcin producing strains were found in 27% (14/52) controls, 48% (30/63) non-a-A, 57% (34/60) a-A and in 53% (34/64) CRC; Fig. 2The Figs. p>0.05.No significant difference in colicin and microcin production was found between patients with current and previous non-a-A, between individuals with current and previous a-A, between patients with current and previous CRC; .Production of colicin and microcin in men and women is shown in Fig. 4.Colicin Ia and E1 are the most frequently produced colicins in patients with colorectal neoplasia. Absolute numbers of these colicins in each group tested are shown in Fig. 5Microcin mH47, microcin mM and microcin mV were the most frequently found microcins across the investigated groups. Microcin mE492 was not produced by any strain.p>0.05.Our study did not confirm any differences in colicin and microcin production between the caecum, the transverse colon and the rectum in any groups of individuals enrolled in the study; No difference in colicin or microcin production in each group of patients with colorectal neoplasia was found between those prepared for a colonoscopy with polyethylene glycol and those who received other agents for bowel preparation.The profile of colicin and microcin produced in an individual with current adenoma or carcinoma on colonoscopy was not dependent on the localisation of the neoplasia. The same or a very similar profile of bacteriocin production between all segments was observed.Our prospective study has shown crucial and novel results in regards to production of colicins and microcins in patients with current and previous colorectal neoplasia. Fundamental differences in bacteriocin production have been confirmed between males and females.S [Escherichia coli and other related gramnegative bacteria from Enterobacteriaceae family [With exception of colicin JS , colicine family , 21–24. e family , a non-se family , 27, a he family . The cyte family , 29–31.E. coli strains [Colicin E1 inhibited growth of ten cell lines, and colicin A of all eleven tested cell lines. The colon carcinoma line HT29 was insensitive to colicin E1. Higher fraction of cells in apopotosis after treatment with colicin A and E1 was observed in this study [E1 colicin belongs to the pore-forming bacteriocins and appears to be a potentially important virulence factor of  strains , 32. Our strains  reported strains . Chumcha strains  evaluateis study . Furtheris study  and diffis study . SubsequEnterobacteriaceae family, are smaller in size (<10 kDa) when compared to colicins. They are produced in situation of nutrient depletion [Microcins, also produced by bacteria from the epletion , 34. Fouepletion .Klebsiella pneumoniae, acts by forming pores [E. coli Nissle 1917 successfully used in clinical practice, produces microcins mM and mH47 [E. coli Nissle 1917 was selectively associated with cancerous cells [E coli Nissle 1917 could become a suitable vector for delivery of mE492 into the tumour [Microcin mM, mH47 and mV were the most frequently found microcins across the investigated groups in our study. On contrary, microcin mE492 was not produced by any strain, neither in any patients with advanced colorectal neoplasia. Microcin E492, produced by ng pores . Microcing pores . Apoptosng pores  have repng pores . Probiotand mH47 , 39. In us cells . As micre tumour , 41.E. coli strains.Bures et al.  studied We are aware, that the bowel cleansing preparation can alter the composition of mucosal adherent microbiota as it was shown by Harrell et al. . TherefoIncreasing production of colicins and microcins with more advanced neoplasia can not be explained by nutrient deficiency as patients even with advanced colorectal adenoma do not suffer from insufficient supply of nutrients.The absence of difference in production of colicins and microcins between those with current and previous colorectal neoplasia is fundamental. It confirms, that production of bacteriocins is not a consequence of neoplasia being present. The observation rather acknowledges the comprehensive effect of mucosal bacteria: their contribution to the development of colorectal neoplasia, but also their probable protective effect through production of bacteriocins.The later onset of increased production of bacteriocins  during the adenoma-carcinoma sequence in men compared to women is also striking. There is no clear explanation for this, and we can only hypothesize that this could play a role among others in the different incidence of CRC in males and females.Worldwide, there is an immense effort put into the research how to prevent development of colorectal neoplasia and how to optimize treatment of an established colorectal carcinoma. We hope, that bacteriocins could become part of these strategies based on their unique characteristics.Strains isolated from large intestinal mucosa in patients with colorectal neoplasia produce colicins and microcins more frequently compared to controls.Bacteriocin production does not differ between patients with current neoplasia and those with a history of colorectal neoplasia.A later onset of increased production of microcins during the adenoma-carcinoma sequence has been observed in males compared to females.Pore-forming colicins Ia and E1 were the most frequently identified colicins. Microcin mM, mH47 and mV belong to the microcins most frequently found across all the groups.Additional file 1. Primers used for the detection of colicin genes


Tinnitus and hearing impairment are prevalent among headache patients. This study aims to investigate the risk of tinnitus, sensorineural hearing impairment, and sudden deafness in patients with non-migraine headache. Participants included 43 294 patients with non-migraine headache (non-migraine headache cohort) and 173 176 patients with no headache of any type (control cohort) frequency-matched with respect to 10-year age interval and sex from the Longitudinal Health Insurance Database 2005 of the Taiwan National Health Insurance Research Database. The mean age of the non-migraine headache cohort was 28.4 ± 14.9 years, and 58.5% of this cohort was male. The incidence rates of tinnitus, sensorineural hearing impairment, and sudden deafness were compared between cohorts using the Kaplan–Meier method with the log-rank test. A Cox proportional hazard model was used to examine the association of tinnitus, sensorineural hearing impairment, and sudden deafness with non-migraine headache, with adjustment for all covariates. The combined risk of either tinnitus, sensorineural hearing impairment, or sudden deafness was higher in the non-migraine headache cohort than in the control cohort . Subgroup analysis showed that patients in the non-migraine headache cohort were at significantly higher risk of developing tinnitus , sensorineural hearing impairment , and sudden deafness  than were controls. In this population-based study, the risks of tinnitus, sensorineural hearing impairment, and sudden deafness were found to be significantly higher in patients with non-migraine headache than in those without headache. Headache is common in people of all ages. Common chronic headache disorders include migraine, tension-type headache (TTH), and medication overuse headache . SecondaCross-sectional studies have shown that several types of headache are associated with tinnitus and hearing impairment. For example, in 71 patients with tinnitus comorbid with headache, 30 suffered from migraine, 15 from TTH, 5 from cluster headache, and 21 from mixed headache . In 1251The relationship between headache and tinnitus or hearing impairment has been described in serial case and cross-sectional studies –7, 10,11This retrospective cohort study used claims data from the Longitudinal Health Insurance Database 2005 (LHID2005) of Taiwan, a subset of the National Health Insurance Research Database (NHIRD) , derivedThis study was approved by the institutional review board of the Dalin Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Taiwan (No. B10202022). Because this study uses only de-identified data, the review board waived the requirement for obtaining informed consent from all patients.To investigate whether non-migraine headache might increase the risk of tinnitus, sensorineural hearing impairment, or sudden deafness or not. We first identified 124 048 non-migraine patients with outpatient claims between 1 January 1996 and 31 December 2012 . From thWe excluded patients who had several conditions before the index date, including schizophrenia (ICD-9 code 295.90); brain lesion(s), including benign brain tumor  and malignant brain tumor ; ischemic cerebrovascular diseases and basilar artery syndrome ; hemorrhagic stroke ; brain concussion or head trauma ; epilepsy/convulsions ; aneurysm or arteriovascular malformation of brain and neck ; anoxic brain damage (ICD-9 code 348.1); brain abscess/encephalitis ; and ear diseases, including vestibular schwannoma (ICD-9 code 225.1), conductive hearing loss (ICD-9 code 389.0), otosclerosis (ICD-9 code 387.9), tinnitus , sensorineural hearing impairment , and sudden deafness (ICD-9 code 388.2). Those with missing or unmatched data and those who had migraine  before or after index the date were excluded. Finally, a total of 43 294 patients with newly diagnosed non-migraine headache were enrolled as the non-migraine headache cohort.Next, we identified 516 448 patients without headache of any type from 1996 to 2012, with exclusion of the above-mentioned pre-existing conditions before the index date for screening as controls. For each patient in the non-migraine headache cohort, 4 control patients were randomly selected, with frequency matching by age and sex. The final matched-control cohort included 173 176 patients. The index date was the date of selection.Both cohorts were followed from the index date until the first diagnosis of tinnitus, sensorineural hearing impairment, sudden deafness, death, or the end of 2012, whichever came first. Death was assumed upon withdrawal of a patient from the NHI program ,15.We identified comorbidities according to ICD-9 codes present before or on the index date, including sleep disorders , heart disease , hypertension (ICD-9 codes 401–405), diabetes (ICD-9 code 250), hyperlipidemia (ICD-9 code 272), chronic kidney disease , and chronic hepatitis . Geographic region of residence  and urbanization level  were also included to minimize potential confounding due to urban–rural differences in accessibility to medical care in Taiwan.6 person-years) with 95% confidence intervals (CIs) for tinnitus, sensorineural hearing impairment, and sudden deafness using the Kaplan–Meier method between cohorts using the log-rank test. After ensuring the assumptions of proportional hazards, we used the Cox proportional hazard model to examine the association of tinnitus, sensorineural hearing impairment, and sudden deafness, combined and individually, with non-migraine headache, with adjustment for all covariates . We analyzed all data using SAS  and SPSS  and considered a 2-sided p <0.05 statistically significant.Pearson’s chi-squared test and Student's t-test were used to compare categorical and continuous variables, respectively, between cohorts. We determined and compared the incidence rates (per 106 person-years of follow-up in the non-migraine headache cohort and 52.9 in the control cohort (p < 0.001). The cumulative incidence of each individual disorder  was significantly higher in the non-migraine headache cohort than in the control cohort (p < 0.001) (The mean duration of follow-up was 7.79 years for the non-migraine headache cohort and 14.49 years for the control cohort (p < 0.001). By the end of follow-up, the percentage of patients who also had tinnitus, sensorineural hearing impairment, or sudden deafness was 9.4%  in the non-migraine headache cohort and 7.7%  in the control cohort (p < 0.0001). The incidence of tinnitus, sensorineural hearing impairment, or sudden deafness was 121.1 per 10< 0.001) .The crude and adjusted hazard ratios (aHRs) for all 3 disorders combined  were 2.85  and 2.73 , respectively . As compThe crude HR and aHRs for each individual disorder as compared to the non-migraine cohort were 3.05  for tinnitus, 1.89  for sensorineural hearing impairment, and 2.14  for sudden deafness in non-migraine headache cohort, respectively .In this large-scale cohort study based on data from the Taiwan NHIRD, we investigated the risks of tinnitus, sensorineural hearing impairment, and sudden deaf associated with non-migraine headache. We found that patients with non-migraine headache are at significantly greater risk of tinnitus, sensorineural hearing impairment, and sudden deafness than are those without chronic headache.TTH and migraine are the most common types of headache among the general population ,4. AlthoThis study excluded migraine and some types of secondary headache in the non-migraine headache cohort and excluded all types of headache in the control cohort. Thus, the non-migraine headache cohort mainly includes patients with TTH, medication overuse headache, temporal or giant cell arteritis, or idiopathic low cerebrospinal fluid pressure . In addiPoor sleep, hypoxia, neural inflammation, and oxidative stress damage are important factors in the pathogenesis of primary headache ,18, tinnThis study has several limitations. Misclassification bias introduced by the use of ICD-9-CM might be present. Second, ‘migraine’ might be recorded as ‘headache’ in the dataset, increasing the rate of false-positive diagnoses in the non-migraine headache group. Thus, our conclusions should be interpreted conservatively. Third, unmeasured variables such as noise exposure or medication use may confound the study results.Taken together with the findings of previous studies, our results suggest that non-migraine headache is associated with increased risks of tinnitus, sensorineural hearing impairment, and sudden deafness. Therefore, clinicians should pay close attention to the history of headache when caring for patients with tinnitus and hearing impairment.


Introduction: Previous studies have proven the effectiveness of bilateral cochlear implantation compared to unilateral cochlear implantation. In many of these studies the unilateral hearing situation was simulated by switching off one of the cochlear implants in bilateral cochlear implant users. In the current study we assess the accuracy of this test method. Does simulated unilateral hearing (switching off one cochlear implant) result in the same outcomes as real life unilateral hearing with one cochlear implant and a non-implanted contralateral ear?Study design: We assessed the outcomes of one arm of a multicenter randomized controlled trial.Methods: In the original trial, 38 postlingually deafened adults were randomly allocated to either simultaneous bilateral cochlear implantation or sequential bilateral cochlear implantation. In the current study we used the data of the sequentially implanted group (n = 19). The primary outcome was speech perception-in-noise from straight ahead. Secondary outcomes were speech perception-in-silence, speech intelligibility-in-noise from spatially separated sources and localization capabilities. A within-subjects design was used to compare the results of hearing with one cochlear implant and a non-implanted contralateral ear (1- and 2-year follow-up) with the results of switching off one cochlear implant after sequential bilateral implantation (3-year follow-up).Results: We found no significant differences on any of the objective outcomes after 1-, 2-, or 3-year follow-up.Conclusion: This study shows that simulating unilateral hearing by switching off one cochlear implant seems a reliable method to compare unilateral and bilateral hearing in bilaterally implanted patients.Clinical Trial Registration: Dutch Trial Register NTR1722. Cochlear implantation (CI) has become a widely applied intervention in the treatment of patients with severe to profound bilateral sensorineural hearing loss, who obtain limited benefit from conventional hearing aids.Although many patients with a single cochlear implant achieve high levels of speech perception-in-silence, even the most successful cochlear implantees experience difficulty with speech perception-in-noise and localization capabilities , 2.In 2009, our study group started a randomized controlled trial (RCT) concerning the effectiveness of simultaneous bilateral CI (simBiCI) compared with either 1) unilateral CI (UCI)  unilater, or (2) Earlier (cohort) studies showed similar benefits of BiCI compared to UCI, however they used different methods to simulate the unilateral listening situation. In most of these studies, differences between bilateral and unilateral hearing were assessed using a within-subjects study design by switching off one cochlear implant in a group of bilaterally implanted patients and comparing the results with the bilateral listening situation –13.Our hypothesis was that this simulated unilateral listening situation would not be representative for an actual UCI situation. The electrode in a patient with bilateral implants would have diminished residual hearing. On the other hand, patients with bilateral implants are used to listening with two ears in everyday life, while in patients with a unilateral cochlear implant, patients may not have used one ear for an extensive period of time.We performed the current study to assess the reliability of switching off one cochlear implant as a method to simulate unilateral hearing in bilateral cochlear implant users.Data for the current study were collected as part of a multicenter RCT that compared simBiCI to seqBiCI –4.This study was approved by the Human Ethics Committees of all participating centers  (NL2466001808), registered in the Dutch Trial Register (NTR1722) and conducted according to the Declaration of Helsinki. Written informed consent was obtained from all participants.All participants eligible for CI were discussed in our cochlear implant team. Inclusion- and exclusion criteria were verified for each participant . After rIn the current study, we will focus on the first implanted side (CI1) in the seqBiCI group, by using a within-subjects design. Patients in this arm received their second cochlear implant 2-years after their first cochlear implant. Objective outcomes were measured after the first implantation at 1- and 2- years follow-up. After 2-years of follow-up, the patients received their second implant. In order to assess whether simulated unilateral hearing (switching off one cochlear implant) provides the same outcomes as real life unilateral hearing, we compared the data of this group 1- year after unilateral implantation (1-year follow-up) with the situation after bilateral implantation (3-year follow-up), in which we switched off the second cochlear implant (CI2). As a sensitivity analysis, to correct for a possible learning effect with the first implanted ear, we also compared the unilateral 2-year follow-up data with the simulated unilateral 3-year data (switching off the second cochlear implant).The primary outcome was speech perception-in-noise from straight ahead, measured with the Utrecht-Sentence Test with Adaptive Randomized Roving levels resulting in a speech reception threshold in noise (SRTn). A lower threshold value reflects better speech perception. A SRTn of 30 dB was considered speech perception in relative silence and was used as a cut-off point for all scores above 30 dB.The other outcomes were (1) speech perception-in-silence, (2) speech intelligibility-in-noise from spatially separated sources (SISSS), and (3) localization capabilities. All these objective tests were conducted using the AB-York Crescent of Sound set-up. In previous articles of our study group more detailed information was presented about the test procedures and setup.Speech perception in silence was measured using the standard Dutch consonant-nucleus-consonant (CNC) test.In the SISSS, in which the outcome is also expressed as an SRTn, sentences were presented from 60° azimuth to the left of the patient and noise from 60° azimuth to the right of the patient (S-60 N+60) or vice versa (S+60 N-60). When sounds come from different directions, participants usually have a best performing situation and a worst performing situation. In current study, in which we evaluate the unilateral group or situation, speech presented to the cochlear implant side was indicated as the best performing situation. If speech was presented to the contralateral ear and noise to the cochlear implant side we indicated this as the worst performing situation.For the localization test, participants were instructed to face the loudspeaker in front of them during the entire procedure. Thirty phrases were presented randomly at 60, 65, or 70 dB SPL from one of the loudspeakers. The results were percentage correct responses in three localization conditions: 15° angle azimuth between 5 loudspeakers, 30° angle azimuth between 5 loudspeakers, and 60° angle azimuth between 3 loudspeakers.All gathered data were double-checked by two independent persons who did not have any further connections to the otorhinolaryngology department.t-test for numeric normally distributed data, the Wilcoxon signed rank test for non-normally distributed data and the chi-square test for ordinal data.In order to compare baseline characteristics, means or medians were reported depending on normality of data. We used the Student Between December 2009 and September 2012, a total of 19 participants were included in the seqBiCI group. During the second and third year, two participants in the seqBiCI group withdrew because of personal reasons. Another participant was excluded because of poor results with the first implant and low expectations after sequential implantation owing to central deafness caused by rhesus antagonism .We found no significant differences between simulated unilateral hearing (switching off one cochlear implant) 1-year after seqBiCI and real life unilateral hearing 1- and 2-years after UCI. The results of all outcomes separately are presented in In order to assess methodological issues with simulation of cochlear implant use, in present study we assessed whether simulated unilateral hearing (switching off one cochlear implant) provides the same outcomes as real life unilateral hearing.Binaural hearing has been proven to be superior to unilateral hearing with regard to speech perception in noise and sound localization , 14–17. We assessed if this is a reliable test method.However, the current study demonstrated similar results after UCI and hearing with one cochlear implant switched off after seqBiCI on speech perception-in-silence, speech intelligibility-in-noise and localization tests.This is the first study that reports whether simulated unilateral hearing (switching off one cochlear implant) provides the same results as real life unilateral hearing.A strength of our study is that we used a prospective within-subjects study design. All data were collected at fixed moments. Secondly, by measuring after a follow-up of at least 1-year after implantation, it was safe to assume that patients were used to their implants and that we had corrected for a possible learning effect. Furthermore, at time of inclusion, all patients suffered from profound sensorineural hearing loss [pure-tone average of greater than 90 dB ]. Because of the profound hearing loss in the contralateral ear (second implanted ear) patients were not used to listening with two ears after the first implantation. Therefore, the situation before the second implantation can be considered as actual unilateral hearing.As this study was based on a secondary analysis from a larger RCT, a power analysis for the present study was not performed and the study may be underpowered.We found no significant differences between simulated unilateral hearing (switching off one cochlear implant) and real life unilateral hearing. This study shows that simulating unilateral hearing by switching off one cochlear implant seems a reliable method to compare unilateral and bilateral hearing in bilaterally implanted patients.This study was carried out in accordance with the recommendations of the medical ethics committee (MEC) of the Academic Medical Center (AMC) with written informed consent from all subjects in accordance with the Declaration of Helsinki. The protocol was approved by the MEC AMC. This study was also registered in the Dutch Trial Register (NTR1722).All authors have made substantial contributions to the paper. AvZ collected parts of the data, analyzed the data, and wrote the paper. YS designed the study, recruited and included participants, collected parts of the data, and critically revised the paper. VK collected parts of the data and critically revised the paper. GvZ provided audiometric advice and critically revised the paper. RS recruited and included participants and critically revised the paper. IS provided advice and was involved in the methodology for present study, statistical analysis and co-authored the paper.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.


In keeping with recent developments in basic research; the importance of the Cathepsins as targets in cancer therapy have taken on increasing importance and given rise to a number of key areas of interest in the clinical setting. In keeping with driving basic research in this area in a translational direction; recent findings have given rise to a number of exciting developments in the areas of cancer diagnosis; prognosis and therapeutic development. As a fast-moving area of research; the focus of this review brings together the latest findings and highlights the translational significance of these developments. Recently, the cathepsin family of proteases have gained significant attention particularly in the field of cancer biology due to their ability to modulate the architecture of the Extracellular Matrix  ,2.Together, the 15 lysosomal proteins that make up the human family of cathepsin proteases, which can be broadly classified into aspartic -, serine - and cysteine -proteases types . AdditioTaken with the observations that some of these proteases are found to show upregulated levels of expression in cancer progression, it comes as no surprise therefore that some of them have been validated as potential therapeutic targets in normal developing cancer types and cancer types that show resistance to regular chemotherapeutic regimes. Of significance are the cathepsins which are over-expressed in cancers that show biological features consistent with late stage cancer and tumor metastases . HoweverAs the expression of certain cathepsins are significantly upregulated in certain cancer types it has also been reported that their ablation has the potential to sensitize cancer cells to conventional chemotherapeutic treatments. For example, this effect has been seen in the instances of Cathepsin L over-expression in ovarian and lung cancer cells which offers a key perspective that can be exploited for the purpose of therapeutic design ,14,15. MBroadly, the cathepsin family range in 50 to 90 KDa in size which after translation are inserted into the endoplasmic reticulum (ER), where they enter the secretory pathway as inactive zymogens. Within the ER, they are post-translationally modified and following their transport through the trans-golgi network, become trans- and auto-activated upon reaching the acidic environment of the endosomes ,17,18. OAll cysteine cathepsins can be regulated by the naturally-occurring endogenous cystatin inhibitors A. This kThe cystatins (or type 2 cystatins) are approximately 115 aa in length, are secreted and the group members characteristically containing two conserved disulphide bonds . The kinHow these inhibitor proteins exert their activity in disease through the modulation of specific cysteine cathepsins  has beenIn this review, we evaluate recent advancements in the use of these various approaches and the potential they hold in quantifying active cathepsin proteases for diagnostic, prognostic and therapeutic purposes.More recently, the transcriptional and post-translational regulation of the cystatin family members have gained greater importance, due to their key regulatory effects on cathepsin protein activity and because they hold good potential in diagnostic and prognostic assay design. Additionally, they have also been found to modulate nuclear gene expression, while at the same time their genes are amenable to transcriptional regulation upon cell cycle arrest. For example, Mori et al. (2016) found cystatin C transcription to be negatively regulated by p53 in response to DNA damage and a lower expression level of cystatin C suggested to offer a poor prognosis in breast cancer (BC) . CystatiCollectively such findings highlight the on-going importance of characterizing cystatin gene and protein expression, clearly as their ability to naturally inhibit aberrant cathepsin activity has implications in the clinical setting. Their input potentially offering greater effectiveness of chemotherapy, particularly in the context of tumor proliferation, chemotherapeutic resistance and tumor invasion.Recently, significant emphasis has been placed on the use of quantified cathepsin (and/or cystatin expression ratios) in developing novel and more accurate diagnostic patient tests. These are aimed at measuring more precisely these expression products from tumor cells in the absence of any possible interference from ECM-derived stromal cells . Such teSubsequently, Liu et al. (2016)  measuredTo compliment such strategies, the cystatin inhibitor proteins have also been incorporated as diagnostic biomarkers in cancer biology. For example, Komura et al. (2017) observed higher cystatin A protein levels in the serum from 36 pancreatic ductal carcinoma (PDAC) patients in comparison to normal patients . AdditioConsequently, attempts have been made to offer a more robust assay which paints an accurate picture of how potentially active specific cathepsin levels may be derived from disease patients. Therefore, recent assays have been developed to simultaneously quantify the levels of serum cathepsin and cystatin expression. Here, Zhang et al. (2017) quantified cathepsin B and cystatin C ratios from 95 patients with CRC using a variety of control samples and observed statistically significantly higher ratios in disease patients in comparison to normal . In a siTo compliment and develop such studies, alternative (yet synergistic) avenues of investigation that appear to be receiving much attention include the development of activity-based probes (ABPs) in theranostic-based assays. While such assays have the potential to measure directly the catalytic activity and subcellular localization of specific cathepsins, they also having huge potential in being able to be adapted for cathepsin targeting and therapeutic delivery. Initially, such ABPs were designed to bind cathepsin proteins in an activity-dependent manner  but moreCollectively, there has been positive progress reported using a variety of methods quantifying cathepsins, expressing their levels as a ratio of cystatin to cathepsin enzyme activity in vivo, and using a variety cohort sizes, samples or approaches see  may offeAs key mediators of cancer progression, the cathepsins of late have also received much attention as potential prognostic markers in developing assays incorporating tumor size, grading, oestrogen receptor expression and lymph node diffusion . Here, aRecently, assays have incorporated cathepsins B, D, G, K, L, V and S quantification using IHC (from BC patients ), RT-PCROne of the most exciting areas in cathepsin research that has enjoyed a flurry of activity of the last 3 years is the development and publication of therapeutics, either directed at the cathepsins or derived from them. A wealth of knowledge has been published in mapping the active sites for the cathepsins and thus given rise to a number of peptides and inhibitors as candidate therapeutics . While iARF-derived peptide fused to a polypeptide containing a cell entry signal as a drug. This showed very promising outcomes as a therapeutic against a variety of cancer cell types. Alternatively, Kramer et al. (2017) designed a highly penetrative ‘DARPin’ to successfully target cathepsin B activity in mouse BC models with a Ki of 35 picomolar [i concentration as low as 10−10 M [In the case of cathepsin B , a catheicomolar . In a no 10−10 M . Moreove 10−10 M . Finally 10−10 M .50 as low as 13.12 μM in the inhibition of BC metastasis [In the case of cathepsin D, small molecule inhibitors against this were used to sensitize triple-positive and -negative BC cells with levels of inhibition exceeding 50% (at 25 M) . Thus, stastasis .Collectively, all studies show great promise in treating cathepsin-mediated cancer to varying degrees and efficacies in model systems and clearly good progress is being made in defining cathepsin-specific inhibitors with enhanced efficacy of action. Moreover, recent studies clearly demonstrate the use to which cathepsin-specific cleavage sequences or cathepsin-derived sequences can be utilized in targeting tumors more effectively. Further studies addressing how effective these are in animal, translational or clinical contexts by modulating conventional chemotherapeutics to be more effective against resistance are eagerly awaited.Based on the articles cited herein, the area of diagnostics holds great promise for designing a cathepsin-based assay that is quick, accurate and reproducible for the diagnostics of gastric, lung, pancreatic bladder, colorectal and esophageal cancers. Simultaneously, such assays may offer great insight into how bio-active secreted or intracellular specific cathepsin proteins may be, particularly in the development of ABPs. As intracellular compartmentalization of some cathepsin and cystatin proteins can potentially affect their observed ratios in serum, there are limitations in using assays that exclusively analyze serum levels alone. Thus, highlighting the importance of incorporating tumor biopsy analysis using IHC (or even cell staining techniques with ABPs) with ELISA assays on sera to ascertain cathepsin/cystatin expression ratios (as shown in From the perspective of therapeutics development, the general findings look very encouraging for targeting extracellular cathepsin proteins and studies have started addressing their efficacy in an approaches that utilizes combined chemotherapeutic treatment in model laboratory systems. This work is indeed at the stage where it can transition into clinical models. For example, how effective such cathepsin therapeutics maybe in a combined therapeutic setting (such as with Tocilizumab  or RituxWhile great strides have been made in the area of cathepsin assay design for diagnostic and prognostic uses, future studies hold great potential. It would appear that at this juncture, we are experiencing a ‘period of transition’ that is being quickly transformed by harnessing the rapidly evolving potential of cathepsin theranostics. When taken together, the future for cystatins and cathepsins in developing cancer research from ‘bench to bedside’ appears to be progressing in good balance and for which the future looks very rewarding.


Objective: Long-term positive energy balance promotes the development of obesity, a main risk factor for type 2 diabetes mellitus (T2DM). While an association between increased resting metabolic rate (RMR) and insulin sensitivity (IS) was shown previously, the underlying mechanisms remain unclear. Aim of the mediator analysis was to investigate the role of inflammation within the association between RMR and IS.Methods: Anthropometric, clinical, and lifestyle data were collected according to standard operating procedures. RMR was measured using indirect calorimetry. Homeostasis model assessment for insulin resistance (HOMA-IR) was used as an IS parameter and C-reactive protein (CRP) was measured to represent the inflammatory status. Statistical analyses were performed using SPSS.Results: The analysis included 782 adults  with a mean age of 32.4 ± 12.0 years and a mean body mass index (BMI) of 24.6 ± 5.2 kg/m2. Regression analysis indicated a significant evidence for associations between RMR and HOMA-IR  and CRP and HOMA-IR  after adjustment for fat-free mass, sex, age, and study site. Results of the mediator analysis did not support the hypothesis that CRP is a mediator for the association between RMR and HOMA-IR. These results did not change after participant stratification according to sex or BMI.Conclusion: A significant evidence for an association between RMR and IS was shown in a large cohort. However, the inflammatory status, determined via CRP levels, was not a mediator within this association. The Global Burden of Disease consortium estimates that the prevalence of obesity and type 2 diabetes mellitus (T2DM) has more than doubled globally in recent decades . Data frBoth obesity and T2DM are characterized by not only impaired insulin sensitivity (IS) but also low-grade inflammation , 5. SubaTo date, the role of the inflammatory status within the association between RMR and IS as well as the impact of inflammation on RMR remains unclear. Therefore, the present analysis was conducted to evaluate the role of the inflammatory status, represented by CRP levels, within the association between RMR and IS. In addition, sex- and body mass index (BMI)-specific subanalyses were performed to study potential confounders.2. Subjects who appeared healthy and showed no history of severe diseases or surgery within the last 3 months and did not have acute physical impairment were eligible. Pregnant and breast-feeding women were excluded. Participants were studied after an overnight fast of about 10–12 h and had to refrain from smoking and physical activity for 12 and 24 h before RMR measurement, respectively. All study procedures were in agreement with the ethical guidelines and were approved by the ethical committee of the Technical University of Munich, Germany (Number 2719/10 S). Written informed consent was obtained from all persons before participation in the study. For the present analysis, participants with overt diabetes, based on self-reporting or with a fasting plasma-equivalent glucose value above 125 mg/dl, were excluded. Furthermore, individuals on medications affecting the sympathetic nervous system such as beta blockers were excluded. For subanalyses, individuals with hypertension (systolic/diastolic blood pressure >140/90 mmHg), defined according to the guidelines of the European Society of Cardiology . Using a validated electronic device  blood pressure was measured as recommended by the American Heart Association  and reported to the nearest 0.1 cm. Body composition was assessed by bioimpedance analysis using Tanita BC 418 MA . Measurements were performed in light clothing and with voided bladder, whereby 1.0 kg was subtracted automatically for clothes. BMI was calculated as the quotient of weight and height squared . Serum collection tubes were stored half an hour at room temperature before centrifugation at 3,000 rpm with 4°C for further 10 min. Serum samples were analyzed for hs-CRP level by turbidimetry  and insulin level by a chemiluminescence immunoassay  by SYNLAB Holding Deutschland GmbH . Homeostasis model assessment for insulin resistance (HOMA-IR) was calculated as follows: Fasting blood samples were obtained after RMR measurement. Fasting blood glucose level was analyzed from whole blood using a HemoCue® Glucose 201μUml)405.2) and carbon dioxide production (VCO2) resulting from substrate oxidation continuously every 5 min under a ventilated hood with the participant fully awake and motionless. RMR was calculated by the following equation . All RMR measurements were performed according to a standard operating procedure. The initial 5 min of the measurement were considered as an acclimatization phase before the actual 30 min measurement. The flow rate was assessed for each participant and adjusted to meet a fraction of expired carbon dioxide of 1.0%, as recommended by the manufacturer. The Quark software recorded oxygen consumption , BMI, insulin, or plasma-equivalent glucose on RMR. These models were further adjusted for age, sex, study site, and FFM. The regression models were also performed in subgroups according to sex. HOMA-IR and CRP values were not normally distributed and were therefore log-transformed for the regression analysis. A mediator analysis, according to Baron and Kenny .Characteristics of study participants, and their clinical parameters, were summarized using means ± standard deviation (SD) and percentages (%). The continuous variables were compared between males and females or between individuals with BMI >24.9 kg/mn = 782, 66.1% females) are shown in Table 2, and mean FFM was 54.5 ± 11.9 kg. Mean systolic and diastolic blood pressure values were 113.8 ± 13.3 and 70.4 ± 8.9 mmHg, respectively. Approximately 4% of the participants (n = 35) had an increased blood pressure (>140/90 mmHg). Most participants (77.0%) were non-smokers. Mean RMR was significantly higher in males compared to females . There were no significant differences in age or BMI between males and females (p > 0.05). Compared to females, males had a significantly higher FFM  but a significantly lower mean fat mass . Mean HOMA-IR was 1.1 ± 0.9 , whereas mean CRP was 0.20 ± 0.35 mg/dl  (Table p ≤ 0.001). Mean HOMA-IR and CRP values were significantly higher in individuals with BMI >24.9 kg/m2 compared to normal-weight subjects (data not shown).Baseline characteristics of the study participants (p ≤ 0.001) after adjustment for age, sex, and study site and between RMR and BMI  after adjustment for age, sex, FFM, and study site. Figure p ≤ 0.001) and females  . A significantly positive association was observed between RMR and insulin levels , but not between RMR and plasma-equivalent glucose levels .Significantly positive associations were observed between RMR and FFM  Figure . As showp ≤ 0.001) and between CRP and HOMA-IR , with RMR as the dependent variable  describing an additional result. The mediator analysis did not support the hypothesis of a mediating effect of CRP on the association between RMR and HOMA-IR in both unadjusted and adjusted analyses since HOMA-IR remained highly significant  after inclusion of CRP  did not show evidence for a mediator role of CRP (Table 2) and a merged group of participants with a BMI of equal to or >25.0 kg/m2 revealed that the association between RMR and HOMA-IR remained significant  only in the group of persons with overweight or obesity  showed no evidence for CRP as a mediator of the HOMA-IR–RMR association (Table No significant evidence for a mediator role of CRP was observed if participants with hypertension (>140/90 mmHg), with dyslipidaemia, with drug intake such as statins and aspirin, or with IFG levels were excluded (data not shown). A separate analysis including only participants with IFG n = 8 showed nThe current analysis of a large cohort of adults with varying BMI confirmed that RMR was positively associated with FFM and BMI. Moreover, a positive association between RMR and HOMA-IR as a measure of IS, as well as CRP as a marker of the inflammatory status was observed. However, the mediator analysis suggested that CRP was not a mediator contributing to the association between RMR and HOMA-IR.The positive association between RMR and HOMA-IR, which was reported in previous studies , 25–28, Another aspect is immunological function which requires up to 15% of daily energy expenditure . In clinObesity, impaired IS or T2DM are characterized by low-grade inflammation frequently assessed by CRP as well as interleukin 6 (IL-6) or tumor necrosis factor alpha (TNFα) , 6, 37. n = 19) represented a significantly higher mean BMI of 32.9 kg/m2 compared to the group with a lower CRP level (p ≤ 0.001). Therefore, BMI might itself play a crucial role in diabetes and the inflammatory status. Lending support to this possibility, subanalysis within the Chennai Urban Rural Epidemiology Study (CURES) found significant correlations between CRP, FM, and HOMA-IR (Nevertheless, the underlying causes of increased RMR in subjects with impaired IS or overt diabetes as well as the impact of an activated immune system are still debated, and the mechanisms are not fully understood. The present analysis showed significant associations between RMR, CRP and HOMA-IR. Therefore, it is of added value to investigate whether CRP mediates the association between RMR and IS, represented by HOMA-IR. However, the results of the present analysis did not reveal significant evidence for CRP as a mediator within the association between RMR and HOMA-IR. One possible explanation for this outcome might be related to the current sample consisting mainly of young and healthy adults, in contrast to other studies investigating the effect of CRP on RMR in cohorts with participants suffering from severe diseases , 35, 36. HOMA-IR . In addi HOMA-IR . This fi HOMA-IR . Further HOMA-IR . From a  HOMA-IR , 42. NonThis is the first analysis investigating the potential role of CRP as a mediator within the association between RMR and HOMA-IR in a large Caucasian population. For the mediator analysis the frequently used methodology of Baron and Kenny  was applRMR was measured by indirect calorimetry which is considered as the gold standard for measuring energy expenditure . In the Moreover, HOMA-IR was used as an indirect parameter of IS. Participants with reported overt diabetes and fasting plasma-equivalent glucose levels above 125 mg/dl were excluded from the dataset. Further established methods to determine the diabetes status such as an oral glucose tolerance test or hemoglobin A1c measurement were not used. As CRP and HOMA-IR were not normally distributed, values were log-transformed for calculation, and the possible interference of the log values cannot be ruled out. In addition, even if the criteria for each stage of a mediator analysis were given it is still unclear whether the non-significant association between RMR and fasting plasma-equivalent glucose values has an impact on the results.The current analysis provided significant evidence for an association between RMR and IS in a large cohort of healthy adults. Conversely, there was no significant evidence for a mediator role for circulating CRP within the association between RMR and HOMA-IR. The findings were not conclusive due to the known low specificity of CRP and likely due to the healthy and young study population. Further studies are needed to replicate these findings and investigate the largely unexplained association between glucose metabolism and energy expenditure.TD and TS performed the measurements. TD was in charge of the statistical analysis of the data. TD and CH processed the experimental data, contributed to the idea, and design of the research, performed the analysis, drafted the manuscript, and designed the figures and tables. Therefore TD and CH share the first authorship. LS gave statistical advice, aided in interpreting the results and worked on the manuscript as a native speaker. JP and TS contributed to the design and implementation of the research and to the writing of the manuscript. HH designed and supervised the research project. All authors discussed the results, commented on the manuscript, and approved the final version to be submitted.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.