Hearing loss is the fourth most common chronic disease, but studies on the relationship between hearing loss and socioeconomic factors are limited. We aimed to examine the association between hearing loss and socioeconomic factors among 35–70 year adults in southwest Iran.This population-based cross-sectional study was conducted in the baseline of Hoveyzeh cohort study in adults aged 35–70 in southwest Iran between 2017 and 2021. Information on socioeconomic factors, demographic characteristics, comorbidities, family history of hearing loss, and noise exposure was collected. We assessed the relationship between three levels of socioeconomic factors  with sensorineural hearing loss (SNHL). Multiple logistic regression was used to adjust the potential confounders.Among a total of 1,365 assessed participants, 485 patients were diagnosed as having hearing loss, and the other 880 individuals were diagnosed without hearing loss, which is considered the case and the control group, respectively. At the individual level of socioeconomic, the odds of having hearing loss in the participants with high school education and diploma, [OR = 0.51 (95%CI:0.28–0.92)], and the individuals with university education [OR = 0.44 (95%CI:0.22–0.87)] were significantly lower than the illiterate participants. At the household socioeconomic level, the odds of having hearing loss were lower for those with poor [OR = 0.63 (95%CI:0.41–0.97)] and moderate [OR = 0.62 (95%CI:0.41–0.94)] wealth status vs. those with the poorest wealth status. In the area level socioeconomic, although the odds of hearing loss in the residents of affluent areas were slightly lower than the residents of deprived areas, there was no significant difference among the groups.The individuals with hearing loss may have insufficient education and income. Hearing loss is the most prevalent sensory disorder in the human population , which hPrevious research has established health disparities in health outcomes based on socioeconomic status (SES) , 7. DespOtologic problems, including hearing loss, have a direct negative effect on people's daily life, and therefore access to care and treatment services is very important. However, in some countries, including Iran, these services are not evenly distributed and are mostly concentrated in large medical centers. In addition, the provision of screening, treatment, rehabilitation, and rehabilitation services for hearing loss is expensive. Therefore, the frequency of hearing loss is expected to differ in various socioeconomic statuses.So far, evidence to evaluate the association of hearing loss with socioeconomic factors has been limited. No research has so far been designed to examine the relationship between sensorineural hearing loss (SNHL) and socioeconomic factors in adults in Iran. At the same time, there is an urgent public health need, and understanding this relationship can effectively provide more appropriate recommendations for hearing loss prevention strategies. Therefore, we designed a cross-sectional study to identify the relationship between hearing loss and the socioeconomic factors at the three levels of individual, household, and area socioeconomic status in 35–70 years adults.This study was a population-based cross-sectional study conducted in the Hoveyzeh Ear Cohort Study (HECS). HECS is Iran's first cohort study to assess ear health and audiology. It was designed in the context of the Hoveyzeh cohort study (HCS) in southwest Iran, which focused on non-communicable diseases. In the HCS enrollment phase, all eligible people aged 35 to 70 in the region were invited, and 85.16%  accepted the invitation and entered the study . We inviThe Ethics Research Committee approved this study of Iran University of Medical Sciences with a code of ethics I.R.IUMS.REC.1399.1441. This study was conducted based on the Helsinki Declaration. Participants signed a written informed consent form for the interview and audiometric evaluation.In this study, we assessed participants‘ socioeconomic status at three levels. Educational levels, resident type, and employment were individual-level socioeconomic indices. Participants' educational levels were categorized as illiterate, primary school, secondary school, high school diploma, and university levels. Residential type is categorized into two groups, rural and urban. Also, the employment variable was defined in the category “Yes” if the participant is employed and “No” if the individual were a housekeeper, retried, or unemployed.The Wealth score was an indicator of SES at the household level. This index was calculated based on the information on household assets, such as motorcycle, car, TV, cell phone, internet access, vacuum cleaner, freezer, washing machine, computer, and household utilities consisting of house ownership and the number of rooms per capita. A principal component analysis (PCA) was performed to assign a coefficient to each asset. Finally, The wealth scores were recorded into quintiles of poorest, poor, moderate, rich, and richest .The Townsend deprivation index is an area-level SES indicator. This index was calculated using four components in each area: the proportion of households without a car, the proportion of non-homeowner households, the percentage of unemployed residents, and the proportion of overcrowded households. Finally, the computed scores were categorized based on the quintiles as five ordered categories; most affluent, affluent, moderate, deprived, and most deprived .We assessed the association between SNHL and demographic variables [sex ], age groups , and having comorbidity . Diabetes is defined as a fasting blood glucose level of 126 mg/dl or higher or using glucose-lowering medications or a self-reported diagnosis of diabetes. Hypertension is defined as having systolic blood pressure above 140 mmHg or diastolic blood pressure above 90 mmHg, using blood pressure-lowering medications, or a self-reported diagnosis of hypertension. Also, hearing risk factors were [a family history of hearing loss (Yes/No) and noise exposure (Yes/No)].A hearing questionnaire was completed for all participants. The information included auditory-vestibular symptoms, noise exposure, and a family history of hearing loss. An experienced audiologist examined the subjects using pure tone audiometry (PTA). Conventional pure tone air conduction (AC) and bone conduction (BC) thresholds are set at octave periods from 250 Hz to 8,000 Hz and 250 Hz to 4,000 Hz, respectively. Normal hearing was determined as the perception of sound stimuli with an intensity of 20 dB HL or less  at all fP < 0.25. All reported p-values were based on two-tailed tests and considered to have a significance level of 0.05. Stata software version 14.0 was utilized for the statistical analysis.Descriptive statistics were performed for quantitative variables by mean, standard deviation, and categorical variables by frequency and percentage. The Chi-square test evaluated the association between hearing loss and demographic variables, socioeconomic status, comorbidity, and hearing factors. We examined the independent associations of demographic, clinic, individual, household, and area-level SES with SNHL using multi-level logistic regression, controlling for confounding factors. In the univariate analysis, the criterion for the initial entry of variables into multiple regression models was n = 485), and those with normal hearing were in the control group (n = 880)] were assessed.In this cross-sectional study, a total of 1,365 participants . The odds of hearing loss in men were 2.7 times great than in women [OR = 2.70 ] ] .P = 0.004), so the odds of hearing loss continuously decreased with education levels  [OR = 0.44 ]. Overall, the odds of having hearing loss decreased with increasing the wealth status of the participants; so it was statistically lower in participants in the poor and moderate quintiles in comparison to the poorest .Although the odds of having hearing loss in the residents of affluent areas were lower than in those living in the deprived areas, after adjusting for confounding factors, this association was not statistically significant .P = 0.006). The diabetic participants had 56 percent more hearing loss than the non-diabetic people [OR = 1.56 ]. The participants with a family history of hearing loss were 2.5 times more likely to have hearing loss than those without a family history [OR =2.66 ] and this association was statistically significant (P < 0.001). Also, a statistically significant association was found between exposure to noise and hearing loss (P < 0.001), so the individual with noise exposure had threefold more odds of hearing loss than the unexposed group [OR =3.09 ]. However, there was no significant association between hypertension and hearing loss (P = 0.295).Moreover, the results showed a significant association between diabetes and hearing loss (We found that education level as an individual socioeconomic level was statistically associated with SNHL, which is an important determinant of hearing loss. At the household level, hearing loss was more prevalent in the poorest households. Besides, at the area level, hearing loss was more prevalent in the most deprived areas, although, there was no statistically significant relationship between SNHL and Townsend Deprived Index. Our findings demonstrated strong associations between hearing loss and sex, age, diabetes, noise exposure, and family history of hearing loss.This study showed a direct relationship between age and hearing loss. Most studies mentioned that age was significantly associated with hearing loss, indicating that older adults were at higher risk than younger adults –20. SomeIn our study, hearing loss was more common in men, even after adjustment for the effects of other variables. Most studies reported men had higher odds of hearing loss , 26, 27.Our result showed that education level was inversely associated with hearing loss. The results of some studies were also in line with our findings , 28–30. We observed an inverse gradient between hearing loss and the wealth index. Participants in lower wealth quintiles experienced significantly high occurrence of hearing loss compared to those in the highest wealth quintile. Perhaps the reason is that in the poorest households, most of their income is spent on daily expenses, including the purchase of food, and the share of income that can be spent on health care is limited.Studies on the relationship between area-level socioeconomic indicators and hearing loss are scarce. Our results demonstrated an inverse but non-significant relationship between the Townsend deprivation index as an area-level socioeconomic indicator and hearing loss so that people who lived in the most deprived areas experienced hearing loss more than those living in the most affluent areas. This finding was consistent with the previous results on the relationship between regional SES and chronic disease . The assMost previous studies have shown that hearing impairment is associated with lower income , 32. LowAccording to several studies, hearing loss in people with type 2 diabetes was more prevalent , 36–38, We showed a statistically significant relationship between hearing loss and a family history of hearing loss. Recently, evidence of gene-environment interactions in adult hearing loss has accumulated . SeveralOur study has several strengths, including conducting this study in the context of a population-based cohort study using a representative sample of the population that can reduce selection bias; examining the relationship of SES at three individual, household, and regional levels separately; applying valid instruments and measure audiometric tests according to the definition of the world health organization.There were some limitations in our study. First, the relationship between hearing loss and noise exposure may be susceptible to recall bias. Hence, the patients with hearing loss are more likely to recall and report a history of noise exposure than the control group. This can overestimate the reported odds ratios. Second, considering the study design, exposure-outcome temporality cannot be demonstrated. Hence, the relationship between hearing loss and literacy may be subject to reverse causality bias.Our findings revealed that SNHL was inversely related to SES. Among the three assessed socioeconomic indicators, education level and wealth index had the association with hearing loss. This shows that individual and household socioeconomic indicators are more associated with hearing loss. Understanding the socioeconomic impact of hearing loss can help health policymakers in planning health improvement programs, including screening and educational interventions. Besides, early diagnosis of hearing loss in high-risk people, treatment and rehabilitation measures were performed earlier and more effectively. The relationship between socioeconomic status and hearing loss is complex, likely involving multiple simultaneous pathways. Therefore, future longitudinal studies will be necessary to better understand the mechanisms behind these associations.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.The studies involving human participants were reviewed and approved by the Vice-Chancellor provided financial support for Research at Iran University of Medical Sciences, Tehran, Iran. The patients/participants provided their written informed consent to participate in this study.MS, NS, BC, PA, and ZR contributed to the design and preparation of the manuscript and its interpretation. ZR was involved in the data collection and perforation. PA, BC, and ZR performed the statistical analysis. All authors have reviewed and approved the manuscript.


CDKL5 gene. Mutations in the CDKL5 gene lead to a lack of CDKL5 protein expression or function and cause numerous clinical features, including early-onset seizures, marked hypotonia, autistic features, gastrointestinal problems, and severe neurodevelopmental impairment. Mouse models of CDD recapitulate several aspects of CDD symptomology, including cognitive impairments, motor deficits, and autistic-like features, and have been useful to dissect the role of CDKL5 in brain development and function. However, our current knowledge of the function of CDKL5 in other organs/tissues besides the brain is still quite limited, reducing the possibility of broad-spectrum interventions. Here, for the first time, we report the presence of cardiac function/structure alterations in heterozygous Cdkl5 +/− female mice. We found a prolonged QT interval  and increased heart rate in Cdkl5 +/− mice. These changes correlate with a marked decrease in parasympathetic activity to the heart and in the expression of the Scn5a and Hcn4 voltage-gated channels. Interestingly, Cdkl5 +/− hearts showed increased fibrosis, altered gap junction organization and connexin-43 expression, mitochondrial dysfunction, and increased ROS production. Together, these findings not only contribute to our understanding of the role of CDKL5 in heart structure/function but also document a novel preclinical phenotype for future therapeutic investigation.CDKL5 (cyclin-dependent kinase-like 5) deficiency disorder (CDD) is a severe neurodevelopmental disease that mostly affects girls, who are heterozygous for mutations in the X-linked  CDKL5 gene, which lead to a lack of CDKL5 protein expression or function [CDKL5 (cyclin-dependent kinase-like 5) deficiency disorder  is a rare and severe X-linked neurodevelopmental disease caused by mutations in the function ,2,3. CDDfunction , affectifunction . Similarfunction .Cdkl5 have been developed to address how CDKL5 dysfunction leads to neurological defects in CDD [Cdkl5 KO mouse models exhibit several behavioral phenotypes that mimic CDD features, such as impaired learning and memory, social interaction, and motor coordination, together with increased stereotypy. Cortical visual response is severely impaired in Cdkl5 KO mice [Cdkl5 KO mice [Cdkl5 KO mice are characterized by accelerated neuronal senescence/death during aging [Cdkl5 KO brain [Several knockout (KO) mouse models for s in CDD ,11,12,13 KO mice , support KO mice ,15,16, a KO mice . Interesng aging  and by ang aging . The finKO brain ,20, impaCDD symptomatology ,21, alonBased on these premises, the overall goal of this study was to carry out the first in-depth investigation into the effect of CDKL5 loss at a cardiac level by exploiting a mouse model of the pathology. Our results demonstrate that heterozygous Cdkl5 deficiency affects cardiac function and cardiac gene expression. We also show that the absence of Cdkl5 has an important implication for mitochondrial integrity/function, promoting oxidative stress and cardiac fibrosis in the adult heart.Cdkl5 is expressed in the mouse heart (Cdkl5 expression was reduced to 39 ± 5.6% in heterozygous (Cdkl5 +/−) female mice in comparison with wild-type (Cdkl5 +/+) mice  mice B and was−/− mice B.Cdkl5 +/−, Cdkl5 −/−, and wild-type (Cdkl5 +/+) mice. Interestingly, we found that both Cdkl5 +/− and Cdkl5 −/− female mice exhibited tachycardia  or decrease (atenolol) in QTc intervals compared to saline  and adrenergic beta 1 (Adrb1) receptors. We found that the mRNA levels of Chrm2 were significantly lower in the hearts of Cdkl5 +/− female mice than in those of wild-type (Cdkl5 +/+) mice  mice I, while e hearts J.Cdkl5 +/− mice, we analyzed the expression of different voltage-gated channels involved in the action potential of cardiac myocytes. In particular, we focused our attention on the genes encoding for the voltage-gated potassium  and sodium (Scn5a) channels. Mutations in these genes are the most common causes of inherited long QT interval syndrome (LQTS), a condition characterized by a prolongation of the QT interval on the ECG, an increase in heart rate, and the risk of ventricular arrhythmias [To investigate the molecular alterations underlying the prolonged QTc interval and elevated heart rate in hythmias .Kcnq1, Kcnh2, or Kcnj2 channels between Cdkl5 +/+ and Cdkl5 +/− female hearts  mice  mice .Hcn4 expression was examined. Interestingly, we found that Hcn4 mRNA levels were significantly lower in Cdkl5 +/− female mice than in wild-type mice  was calculated. No significant differences in heart weight, body weight, or HW/BW were found between Cdkl5 +/− and Cdkl5 +/+ mice (Cdkl5 +/+) and Cdkl5 +/− mouse hearts.To determine whether the absence of +/+ mice . Similar+/+ mice  and the Cdkl5 +/− mice compared with wild-type (Cdkl5 +/+) mice (Col1a2) and the α-3 chain of collagen type 2 (Col3a1)  mice B. Increa(Col3a1) C.Cdkl5 +/− mice in comparison with wild-type (Cdkl5 +/+) mice  mice D.The electrical conduction in cardiac muscle relies on efficient gap-junction-mediated intercellular communication between cardiomyocytes, which involves rapid anisotropic impulse propagation through connexin (Cx)-containing channels, namely Cx43, the most abundant Cx in the heart.Cdkl5 +/− mice in comparison with Cdkl5 +/+ mice  and in the presence of ADP . Glutamate–malate and succinate donate electrons to the respiratory chain at complex I and complex II, respectively. State 3 of respiration, in the presence of glutamate-malate or succinate, was significantly decreased in cardiac mitochondria from Cdkl5 +/− compared to wild-type mice  in ype mice C. In acc+/+ mice D. The coificance D.Cdkl5 +/− mice. We found no significant difference in the density of mitochondria in the myocytes (data not shown), nor was there any difference in the overall size of individual mitochondria , an end product of lipid peroxidation, consistent with a condition of oxidative stress  polymerase 1 (PARP1) and the nuclear factor erythroid 2-related factor 2 (Nrf2), that are known to be activated due to oxidative stress showed increased levels in the hearts of +/+ mice C–F. Simi+/+ mice G,H, whicCdkl5 KO (Cdkl5 +/−) female mouse, exhibits cardiac functional and structural abnormalities. Cdkl5 +/− mice exhibited QTc prolongation and increased heart rate accompanied by impaired cardiac autonomic control. Moreover, the Cdkl5 +/− heart shows typical signs of heart aging, including increased fibrosis, mitochondrial dysfunctions, and increased ROS production.CDD is a very severe and debilitating neurodevelopmental infantile disorder with harsh neurological symptoms such as intractable seizures, neurodevelopmental delay, and autistic-like features. Nevertheless, the improvement of the clinical overview of CDD in the past few years has defined a more detailed phenotypic spectrum; this includes very common alterations in peripheral organ and tissue function, such as gastrointestinal problems, irregular breathing, hypotonia, and scoliosis , suggestCdkl5 +/− mice showed statistically significant increases in heart rate and in rate-corrected QTc intervals, which are indicative of delayed ventricular depolarization and repolarization. This is in line with a recent finding showing that in a cohort of individuals with CDD, the incidence of prolonged QTc or other abnormalities including sinus tachycardia was higher than the prevalence seen within the general population [Cdkl5 +/− mice, homozygous Cdkl5 −/− female mice exhibited tachycardia and a longer QTc interval in comparison with Cdkl5 +/+ mice. Interestingly, the magnitude of the cardiac defects was similar in the two Cdkl5-deficient conditions. This is not surprising, since it has been shown that heterozygous Cdkl5 +/− mice develop some behavioral abnormalities that are comparable to defects identified in homozygous Cdkl5 −/− females [Cdkl5 KO female mice.ECG measurements from pulation . Howeverpulation . Here, u females . The putCdkl5 +/− mice. By applying validated indices of spontaneous RR variability in mice, we assessed cardiac vagal modulation during sleep under baseline conditions and following pharmacological manipulations. Our findings that in Cdkl5 +/− mice, pNN8 and RMSSD indices were lower than in control mice indicate an impairment in the vagal modulation of the heart period in Cdkl5 +/− mice. Compared to saline infusion, treatment with muscarinic antagonist (atropine) did not significantly modify the heart rate or QTc interval in Cdkl5 +/− mice, confirming the reduction in parasympathetic tone in the heart. A growing body of evidence indicates that reduced cardiac parasympathetic activity is a common alteration in brain disorders; it is present in children with autism and Rett syndrome [2 receptor expression in the heart of Cdkl5 +/− mice may be one of the possible mechanisms underlying the parasympathetic dysfunction in the mouse model of CDD. However, since, in many situations, loss of Cdkl5 function appears to lead to impaired neuronal activity [Cdkl5 +/− hearts responded to atenolol infusion by increasing RR and decreasing QTc intervals, indicating the presence of a preserved sympathetic modulation in the Cdkl5 +/− heart. Interestingly, after atenolol infusion, RR and QTc intervals no longer differed between genotypes  are altered in their expression in Cdkl5 +/− female hearts indicates that CDKL5, either directly or indirectly, regulates the expression of genes that play a role in the cardiac conduction systems at the cardiomyocyte level. Loss-of-function mutations in the SCN5A gene, which encodes the α subunit of the cardiac voltage-gated Na+ channel NaV1.5, underlie cardiac disorders, including long QT syndrome [Scn5a genes have been reported in Mecp2-null mice [Hcn4 found in Cdkl5 +/− hearts could also contribute to the altered heart rate in Cdkl5 +/− mice. Our results suggest that Cdkl5 deficiency selectively affects the expression of voltage-gated channel genes in the heart and that abnormal cardiac gene expression may be an arrhythmogenic substrate in Cdkl5 +/− mice.The discovery that two cardiac-specific genes  that cause damage to cellular macromolecules; accumulation of this damage leads to the physiological compromise seen in aging [One of the main risk factors for cardiovascular diseases is aging . Previou decline . Here, win aging . Currentin aging .Cdkl5 +/− hearts when ADP was supplied as a substrate for the ATP synthase (state 3 respiration) and diminished specific activity of complexes I and II. These data are consistent with the decreased level of ATP and the decreased ATP/ADP ratio measured in heart tissue homogenates from Cdkl5 +/− mice compared with wild-type animals. In addition, mitochondria isolated from the hearts of Cdkl5 +/− mice showed increased ROS and lipid peroxidation biomarker (MDA) production compared with controls, suggesting a link between mitochondrial dysfunction and the onset of oxidative and energetic stress.In this study, we found a decrease in mitochondrial oxygen consumption rate in isolated mitochondria from Cdkl5 KO mice identified brain mitochondrial functional abnormalities, including reduced activity of mitochondrial respiratory chain complexes and impairment in mitochondrial ATP production rate [Cdkl5 +/− heart. The presence of mitochondrial paracrystalline inclusions in a clinically characterized group of patients with genetically defined mitochondrial disease [Cdkl5 +/− mitochondria. However, we cannot exclude the possibility that the crystalline-like inclusions represent compensatory responses to mitochondrial stress.These findings are in line with recent evidence that mitochondrial dysfunction and oxidative stress occur in CDD ,56. Cytoion rate ,60, as wion rate ,62. It i disease ,64 suggeCdkl5 +/− female mice could be a consequence of an increase in the generation of ROS due to aberrant mitochondrial function. Similarly, increased LC3-II levels, which are associated with either enhanced autophagosome synthesis or reduced autophagosome turnover [Cdkl5 +/− heart, suggesting an increasing number of autophagosomes.Recent evidence has described a potential role of AKT and autophagy, molecular mechanisms of cellular senescence, in aging-associated organ deterioration. When cardiomyocytes incur oxidative stress, they activate the PI3K/AKT signaling pathway , suggestturnover , may be turnover . We founCdkl5 +/− cardiac myocyte dysfunctions, we found alterations in AKT activity but not in ERK1/2 signaling. Activation of ERK1/2 is generally associated with cell growth and survival, and studies of transgenic mice have shown that selective activation of the ERK1/2 cascade in the myocardium induces adaptive cardiac hypertrophy [Cdkl5 +/− heart, this might explain the lack of ERK1/2 dysregulation in these mice.The principal growth-promoting intracellular signaling pathways that are activated by ROS in cardiac myocytes include not only the AKT pathway but also the mitogen-activated protein kinase cascades (ERK1/2 pathway) . In assoertrophy . Furtherertrophy . Since nCdkl5 +/− heart. We hypothesized that increased levels of Nrf2 are triggered by ROS production and that this increase is most likely a compensatory effort to increase antioxidant defenses in the Cdkl5 +/− context. This should correlate with a reduction in oxidative stress in Cdkl5 +/− hearts; however, this is not the case, and the issue requires further investigation. It is worth noting that PARP activation has been demonstrated to impair mitochondrial function [Cdkl5 +/− heart.Considering the protective molecular pathways that might be triggered by oxidative stress  and autofunction  and promfunction . TherefoCdkl5 +/− heart. However, increasing evidence suggests that ROS production is associated with cardiac arrhythmias; in particular, elevated cellular ROS can cause alterations of the cardiac ion channels, changes in mitochondrial function, and gap junction remodeling, leading to arrhythmic conditions [Cdkl5-deficient heart. Future studies aimed at rescuing the cardiac phenotype in Cdkl5 KO mice, targeting one of the observed defects, could help to shed light on the primary causes that drive cardiac abnormalities in the absence of Cdkl5.At present, we have no evidence for which of the observed Cdkl5-related alterations is the primary cause underlying the structural and functional abnormalities in the nditions ,76,77. Tnditions ,79,80, mCdkl5 −/Y strain in the C57BL/6N background developed in [Cdkl5 +/− and homozygous Cdkl5 −/− females were produced and genotyped as previously described [Cdkl5 +/+ littermate controls were used for all experiments. The day of birth was designated as postnatal day (P) zero, and animals of 24 h of age were considered as 1-day-old animals (P1). After weaning (P21-23), mice were housed three to five per cage with a 12 h light/dark cycle in a temperature- and humidity-controlled environment with food and water provided ad libitum. The animals’ health and comfort were controlled by the veterinary service. Experiments were carried out on a total of 121 adult (3–4-month-old) Cdkl5 KO mice . The study protocols complied with EU Directive 2010/63/EU and with Italian law  and were approved by the Italian Ministry of Health (protocol n° 535/2022-PR). All efforts were made to minimize animal suffering and to keep the number of animals used to a minimum.The mice used in this work were derived from the loped in  and backescribed , and ageCdkl5 +/+, 10 Cdkl5 +/−, and 8 Cdkl5 −/− female mice were instrumented with electroencephalographic (EEG), electromyographic (EMG), and electrocardiographic (ECG) electrodes to characterize the electrical activity of the heart during sleep under baseline conditions. A second group of mice  instrumented with the same surgical protocol also underwent surgery to implant an intraperitoneal (IP) catheter for the continuous infusion of autonomic blockers with the purpose of testing the autonomic modulation of heart rhythm.Eight 2, inhalation route) and treated with intraoperative analgesia  and postoperative antibiotic prophylaxis . For the recording of the EEG signal, mice were implanted with two miniature stainless-steel screws  in contact with the dura mater ; for the recording of the EMG signal, mice were implanted with two multistranded PFA-coated stainless-steel wires  inserted into the nuchal muscles. For ECG recording, two PFA-coated stainless-steel wires were inserted subcutaneously; one was put in contact with muscles in the right-upper quadrant of the thorax, while the second touched the abdominal muscles on the left flank. All electrode wires were then collected in a socket placed over the mouse’s head and fixed with dental cement , Italy) and dental acrylic , Italy). The second batch of mice was also implanted with a silicone catheter [All mice were deeply anesthetized with isoflurane .Mice included in the protocol for testing of the autonomic control of heart rhythm were allowed to recover from surgery and to habituate to the recording settings for 14 days. Each mouse underwent 3 recording sessions (7 h each starting at light on) with acquisition of EEG, EMG, and ECG signals while being continuously infused with either saline, with the muscarinic receptor antagonist atropine methyl nitrate  to block parasympathetic activity to the heart, or with the selective β1-adrenergic receptor antagonist atenolol  to block the sympathetic activity to the heart. Each mouse first received saline infusion and was then randomly subjected to atropine and atenolol infusions. Each infusion was performed at least 48 h after the preceding infusion. The IP catheter was connected to a remote infusion pump  by an external tube prefilled with saline solution or drug solution, as previously described ,82. A raThe EEG signal was cut with a band pass filter between 0.3 and 100 Hz and stored at 128 Hz. The EMG signal was filtered between 100 and 1000 Hz and stored at 128 Hz, while the ECG was filtered between 10 and 1000 Hz and stored at 2048 Hz. Data acquisition was performed with LabVIEW 8.0 software .The EEG and EMG signals were imported and analyzed to automatically score the wake–sleep states  and non-rapid-eye-movement sleep (NREMS)). On the contrary, the ECG signal was imported in Labchart 8.0 , a specific notch (50 Hz) filter was digitally applied to exclude electrical noise, and the signal was then analyzed with the ECG analysis module to automatically detect QRS complexes and their intervals. For each mouse, ECG analysis was restricted to 8 episodes of NREMS and 8 episodes of REMS (longer than 30 s) homogeneously distributed throughout the whole recording. For each session, we calculated the following ECG parameters: P-wave duration, QRS, RR, PR, JT, and QT intervals. To compensate for the elevated mouse heart rate, we corrected the QT interval (QTc) with Hodges’ formula .The vagal (parasympathetic) contribution to cardiac modulation was examined using validated mouse indices computed in the time domain on spontaneous RR fluctuations : pNN8, %Cdkl5 −/−, Cdkl5 +/−, and Cdkl5 +/+ female mice aged 3–4 months were weighed and put under deep anesthesia through inhalation of 2% isoflurane in pure oxygen and sacrificed through cervical dislocation. Hearts were quickly removed, cleaned from the surrounding structures, and thoroughly washed in PBS to remove all blood, then weighed. The ratio of heart weight to body weight (HW/BW) was then calculated by dividing the weight of the heart by the weight of the whole animal. The atrioventricular distance was measured. All measurements were performed by the same person with the same precision scales. Hearts were quickly frozen in isopentane, cooled in liquid nitrogen, and stored at −80 °C until used for RT-qPCR, immunohistochemistry, and Western blot analyses.Adult Cdkl5 −/−, Cdkl5 +/−, and Cdkl5 +/+ female mice and on frozen cortices of Cdkl5 +/+ female mice. Total RNA was isolated using the TRI reagent method , and cDNA synthesis was achieved with 5 μg of total RNA using an iScript™ advanced cDNA synthesis kit  according to the manufacturer’s instructions. Real-time PCR was performed using SsoAdvanced Universal SYBR Green Supermix (Bio-Rad) in a CFX real-time PCR detection system (Bio-Rad). We used primer pairs (Actb and Gapdh) were used as a normalization factor in the RT-qPCR analysis, and relative quantification was performed using the ΔΔCt method.RNA isolation and RT-qPCR were conducted on frozen hearts of er pairs  that proFor histological and immunohistochemistry procedures, frozen hearts were cut with a cryostat (Histo-Line Laboratories) into 7 μm thick sections. Sections were mounted on super frost slides.Sections were stained with hematoxylin and eosin (H&E), immersed in graded alcohols followed by xylene, and mounted in mounting medium .Sections were stained with Masson’s trichrome (MT) using Masson’s Trichrome Stain Kit-Mallory’s ; the procedure was carried out according to the protocol included with the kit.Immunohistochemistry was performed on 7 μm thick frozen sections fixed via immersion in 4% paraformaldehyde  and permeabilized with 0.2% TritonX-100 in PBS. Furthermore, 2% BSA in PBS was used as a blocking reagent. Sections were incubated overnight with anti-β-Catenin antibody or double-stained with anti-connexin 43 and anti-actinin antibodies, washed with PBS, and subsequently incubated for 2 h at room temperature with FITC-conjugated or CY3-conjugated secondary antibodies. The primary and secondary antibodies used in this study are listed in Fluorescence images were taken with an Eclipse TE 2000-S microscope equipped with a DS-Qi2 digital SLR camera (Nikon Instruments Inc.). A light microscope (Leica Mycrosystems) equipped with a motorized stage and focus control system and a color digital camera  were used to take brightfield images of Masson’s trichrome and hematoxylin eosin-stained sections.2. The intensity of β-catenin staining within each area was then quantified by determining the sum intensity of all positive (bright) pixels within the area. Approximately 100 intercalated discs were analyzed from each sample.Starting from 20× magnification images of β-catenin-stained ventricular slices, the area of β-catenin staining in intercalated discs was manually drawn using the Image Pro Plus measurement function and expressed in μmFor the preparation of protein extracts, ventricles were homogenized in RIPA buffer and quantified using the Bradford method as previously described .® TGX™ Gel (Bio-Rad) and transferred to a Hybond ECL nitrocellulose membrane . The primary and secondary antibodies used are listed in TM software (Bio-Rad), which automatically highlights any saturated pixels of the Western blot images in red. Images acquired with exposition times that generated protein signals outside of a linear range were not considered for quantification.Equivalent amounts (50 μg) of protein were subjected to electrophoresis on a 4–12% Mini PROTEANFor the determination of adenine nucleotides, mouse hearts were quickly removed and placed in ice-cold PBS supplemented with 10 mM EDTA. The tissue was homogenized using an Ultra-Turrax (1 cycle for 10 s); then, the resuspension was transferred to a glass potter with a Teflon pestle operated at 1600 r.p.m (10 strokes). All steps were performed at 4 °C. ATP and ADP were extracted and detected as previously described  using HP2, 2 mM Pi, 20 µM EGTA and 0.25 M sucrose, pH 7.4) in a final volume of 1600 µL. Respiratory substrates  were added after signal stabilization. State 4 respiration was recorded for 2 min, and state 3 respiration was induced by the addition of 300 µM ADP. The respiratory rates were expressed in nmol oxygen/min/mg of protein.Intact mitochondria from mice hearts were isolated as previously described . BrieflyCdkl5 +/− and Cdkl5 +/+ mice as previously described [−1) oxidation at λ = 340 nm and 37 °C using a Jasco-v750 spectrophotometer equipped with a cuvette stirring device and thermostatic control. Briefly, the isolated mitochondria were subjected to three freeze–thaw cycles in hypotonic buffer (20 mM phosphate buffer (pH 7.5)) to avoid compartmentalization. Then, 50 µg of mitochondria was added to 700 µL of distilled water in a 1 mL quartz cuvette for 1 min. Then, 100 μL of potassium phosphate buffer , 60 μL of fatty-acid-free BSA (50 mg ml−1), 30 μL of KCN (10 mM), and 10 μL of NADH (10 mM) were added. The volume was adjusted to 994 µL with distilled water. In parallel, a separate cuvette containing the same quantity of reagents and sample with the addition of 10 μL of 1 mM rotenone was prepared. The reaction was started by adding 6 µL of 10 mM CoQ1 -1,4-benzoquinone, Sigma-Aldrich), and the decrease in absorbance at 340 nm was followed for 3 min. Specific complex I activity was obtained by subtracting the rotenone-insensitive activity and normalized to mitochondrial protein content as determined by a Lowry assay [−1) at 37 °C. Briefly, 60 µg of cell lysate, 50 μL of potassium phosphate buffer , 20 μL of fatty-acid-free BSA (50 mg ml−1), 30 μL of 10mM KCN, 50 μL of 400 mM succinate, and 145 µL of DCPIP (0.015% (wt/vol) were added to a 1 mL glass cuvette, and the volume was adjusted to 996 µL with distilled water. After ten minutes of incubation inside the spectrophotometer at 37 °C, the reaction was started by adding 4 μL of 12.5 mM decylubiquinone , and the absorbance decrease at λ = 600 nm was followed for 3 min. To check the specificity of complex II activity, 10 µL of 1 mM carboxine  was added before starting the reaction. The specific activity of complex II was obtained by subtracting the TTFA-insensitive activity and normalized to mitochondrial protein content determined by a Lowry assay [The activity of mitochondrial complexes I and II was assessed in heart mitochondria isolated from escribed  with minry assay . Complexry assay .Cdkl5 +/− and Cdkl5 +/+ female mice using a fluorogenic probe  as previously described [ex = 485 nm; λem = 535 nm) was recorded 10 min after energization using a plate reader .Reactive oxygen species production was assessed in cardiac mitochondria from escribed . For ROSCdkl5 +/− and Cdkl5 +/+ female mice was assessed by measuring the biomarker malondialdehyde (MDA) as previously described by Reilly et al. [Lipid peroxidation in isolated mitochondria from y et al. . BrieflyFor ultrastructural characterization, small heart specimens were fixed in 2.5% glutaraldehyde in 0.1 M cacodylate buffer and post-fixed in osmium tetroxide 1% in the same buffer. After dehydration in graded ethanol, specimens were embedded in araldite. Thin sections were counterstained with uranyl acetate and lead citrate and observed under a Philips CM100 transmission electron microscope .t-test or one-way and two-way ANOVA, followed by Fisher’s LSD post hoc test, as specified in the figure legends. For the in vivo recordings under the baseline condition, data were analyzed with 2-way repeated-measure ANOVAs (considering sleep state and genotype as factors) and corrected for multiple comparisons with Sidak’s test. For mice included in the pharmacological testing of autonomic cardiac contribution, we first calculated the % modification of RR and QTc values induced by the drug compared to saline infusion. Then, for each experimental group, we performed a one-sample t-test, checking for significant deviations from the reference value (100%). A probability level of p < 0.05 was considered to statistically significant. The confidence level was taken as 95%.Statistical analysis was performed using GraphPad Prism (version 9). Values are expressed as means ± standard error (SEM). The significance of results was obtained using a two-tailed Student’s Cdkl5 deficiency impacts heart structure and function in mice and that Cdkl5 +/− mice could be a precious asset to accelerate the comprehension of these aspects. As our work does not provide direct causal evidence linking local CDKL5 expression in the heart or deregulation of direct CDKL5 phosphorylation targets to the electrical activity and structure of the cardiomyocyte, further in vitro and in vivo studies of the effects of specific knockout/rescuing CDKL5 expression in cardiomyocytes are needed. However, the results obtained in the present study could have a considerable impact in promoting a more in-depth investigation of cardiac function in patients for a better understanding of the CDD phenotypes. Although it is known that protein kinases can control cellular signaling by interacting with many targets, our knowledge of the CDKL5 phosphorylation targets is still very limited, hindering the investigation of direct CDKL5-dependent mechanisms underlying cardiac alterations in Cdkl5 +/− mice. However, many alterations found in the heart of Cdkl5 +/− mice are also present in the RTT mouse model, Mecp2-null mice [In conclusion, the present study shows that ull mice ,95,96, aull mice ,98,99, sull mice ,101 suggull mice ,103, the


Retinal implants use electrical stimulation to elicit flashes of light (“phosphenes”). Single-electrode phosphene shape has been shown to vary systematically with stimulus amplitude and frequency as well as the retinal location of the stimulating electrode, due to incidental activation of passing nerve fiber bundles. However, this knowledge has yet to be extended to paired-electrode stimulation.We retrospectively analyzed 4402 phosphene drawings made by three blind subjects implanted with an Argus II Retinal Prosthesis. Phosphene shape  and number of perceived phosphenes were averaged across trials and correlated with the corresponding single-electrode parameters. In addition, the number of phosphenes was correlated with stimulus amplitude and neuroanatomical parameters: electrode-retina (“height”) and electrode-fovea distance (“eccentricity”) as well as the electrode-electrode distance to (“between-axon”) and along axon bundles . Statistical analyses were conducted using linear regression and partial correlation analysis.Simple regression revealed that each paired-electrode shape descriptor could be predicted by the sum of the two corresponding single-electrode shape descriptors The shape of phosphenes elicited by paired-electrode stimulation was well predicted by the shape of their corresponding single-electrode phosphenes, suggesting that two-point perception can be expressed as the linear summation of single-point perception. We also found that the number of perceived phosphenes increased with the between-axon distance of the two electrodes, providing further evidence in support of the axon map model for epiretinal stimulation. These findings contribute to the growing literature on phosphene perception and have important implications for the design of future retinal prostheses. AlthougRetinal prostheses typically acquire visual input via an external camera, which is then translated into electrical pulses sent to a microstimulator implanted in the eye . The stiA series of papers demonstrated that phosphenes elicited by stimulating a single Argus II electrode have a distinctive shape that is relatively consistent over time . PhospheHowever, less is known about how phosphenes combine when multiple electrodes are stimulated. Early research suggested that repeated paired stimulation resulted in reproducible phosphenes as subjects perceived “similar” phosphenes on 66% of trials . But morHere we aim to study the consistency and predictability of the (presumably) fundamental building blocks of form vision: the percepts elicited by single- and paired-electrode stimulation. While single-electrode stimulation is relatively well understood , it rema2.2.1.clinicaltrials.govNCT00407602; completed). All psychophysical experiments were carried out at least six months after device implementation. The study was approved by the Institutional Review Board (IRB) at each subject’s clinical site and was conducted under the tenets of the Declaration of Helsinki. Informed consent was obtained from the participants after explanation of the nature and possible consequences of the study.This study involved three blind participants  with severe RP, ranging from 41 to 70 years in age at implantation . SubjectDue to their geographic location, the participants were not directly examined by the authors of this study. Instead, initial experimental procedures were sent to the clinical site, and trained field clinical engineers performed the experiments as specified. Raw collected data was then sent to the authors for subsequent analysis.2.2.camera mode).Argus II consists of a 6 × 10 grid of platinum disc electrodes, each 200 μm in diameter, subtending 0.7° of visual angle . Electrodirect stimulation mode. Stimuli were charge-balanced, cathodic-first, square-wave pulse trains with 0.45 ms phase duration and 250 ms total stimulus duration. Stimulus amplitudes, frequencies, and the number of stimulated electrodes varied based on the design of each experiment. Stimuli were programmed in Matlab using custom software, and pulse train parameters  to be stimulated, current amplitude, pulse width, inter-pulse interval, and overall stimulus duration) were sent directly to the VPU, which then sent the stimulus commands to the internal portion of the implant using an inductive coil link. The implanted receiver wirelessly received these data and sent the signals to the electrode array via a small cable.All stimuli described in this study were presented in 2.3.Perceptual thresholds for individual electrodes were measured using an adaptive yes/no procedure implemented using custom software see .Participants were asked to perform a drawing task upon electrical stimulation of the retina. Subjects were comfortably seated in front of a touchscreen monitor whose center was horizontally aligned with the subject’s head. The distance between the subject’s eyes and the monitor was 83.8 cm for Subject 1, 76.2 cm for Subject 2, and 77.5 cm for Subject 3.Each stimulus was presented in 5–10 trials randomly amongst other stimuli with different frequency and/or amplitude levels. The stimulus frequency ranged from 6 Hz to 120 Hz, and the amplitude was between 1.25 times threshold to 7.5 times threshold. Within each trial, either one or two electrodes were randomly selected and stimulated; if two electrodes were selected, they were stimulated simultaneously. After delivering each stimulus and before moving to the subsequent trial, subjects were asked to trace the perceived shape on the touchscreen monitor. Shapes were closed by automatically connecting the first and last tracked fingertip location, after which a flood-fill was applied. The drawing data was recorded and converted into a binary shape data file using Matlab, and stored for future analysis. All psychophysical experiments were carried out by local field clinical engineers at each participating site, and the results were forwarded to the authors.Since the validity and reliability of the experiment relied on the ability of our subjects to accurately draw the perceived phosphenes, a control task was conducted where subjects were asked to feel six different tactile shapes made of felt with a cardboard background, and then draw them on a touchscreen . As the 2.4.image moments. For an Phosphene shape was quantified using four parameter-free shape descriptors commonly used in image processing: area, perimeter, major axis length, and minor axis length . An examRaw image moments were used to compute area Phosphene major/minor axis lengths were calculated from the covariance matrix of the phosphene drawing:Phosphene perimeter was calculated using an algorithm described in Phosphene orientation was previously shown to depend mostly on the retinal location of the stimulating electrode  and was If a drawing had more than one phosphene, each shape descriptor was extracted for each perceived phosphene, summed over all phosphenes within a drawing (to account for a variable number of elicited phosphenes), and then averaged across trials.2.5.pulse2percept software  or Topcon 3D-OCT 1000 (Topcon Inc). The SD-OCT scans were obtained 6 months after Subjects 1’s and 2’s implantation and 13 months after Subject 3’s implantation.When performing OCT scanning, the opaque metal electrodes prevent image acquisition directly underneath the corresponding electrode. However, based on the length of the shadow between the electrode and the retinal surface, it is possible to estimate the electrode-retina distance of that electrode . A singlDetails about each subject’s estimated electrode-fovea distances and electrode-retina distances are given in 2.7.To make the collected phosphene drawings amenable to automated image analysis, we manually inspected all 4402 drawings to make sure that:all drawn contour lines were closed ;small specs and other artifacts  were not counted as additional phosphenes.This procedure is explained in detail in 2.8.https://scikit-image.org) was used for calculating different phosphene shape properties, and matplotlib  was used for presenting phosphene drawings and analysis plots.Data entry and statistical analyses were performed in Python . Python package scikit-image . Mean images were then centered over the corresponding electrode in a schematic of the subject’s implant to reveal the rich repertoire of elicited percepts across electrodes.Whereas Subject 1 mostly drew blobs and wedges, which grew larger as the stimulus amplitude was increased, Subject 2 reported seeing exclusively lines and arcs, which got longer with increasing amplitude. The effect of amplitude on phosphene shape was most apparent for Subject 3, where phosphenes that appeared as lines and arcs near threshold turned into blobs and wedges as amplitude was increased.First reported by 3.2.When two electrodes were stimulated simultaneously, participants reported seeing a single phosphene on 53.8% of trials, two phosphenes on 42.8% of trials, and three or more phosphenes on the remaining trials. Three or more phosphenes were generally encountered when single-electrode stimulation itself produced more than one phosphene. Representative examples of phosphene drawings for different electrode pairs are shown in left), their shape resembled the linear summation of the phosphenes reported during single-electrode stimulation. For instance, as shown in Row 1 of the left panel in When paired-electrode stimulation produced two distinct phosphenes , left, tright), the phosphenes reported during single-electrode stimulation appeared to merge into a unified shape. For instance, as shown in Row 2 of the right panel in When paired-electrode stimulation produced a single phosphene , right, Naturally, we asked to what extent the phosphene shape elicited by paired-electrode stimulation could be predicted by the phosphene shape elicited during single-electrode stimulation. To answer this question, we conducted a simple linear regression , where e3.3.To more systematically investigate how different stimulus and anatomical parameters affect phosphene shape, we considered how the four shape descriptors  and distance to the fovea . We found that electrode-As mentioned above, approximately 13% of single-electrode trials elicited multiple phosphenes. Multiple regression revealed that subjects were more likely to see multiple phosphenes as stimulus amplitude increased  increased . This fiIn contrast to In addition, we found that electrode-fovea distance  increased all four considered shape descriptors  machine learning models . In addi


Despite the clear distinction between cortical (cTECs) and medullary thymic epithelial cells (mTECs) in physiology, the cell of origin of thymic carcinomas (TCs) and other thymic epithelial tumors remained enigmatic. We addressed this issue by focusing on AIRE, an mTEC‐specific transcriptional regulator that is required for immunological self‐tolerance. We found that a large proportion of TCs expressed AIRE with typical nuclear dot morphology by immunohistochemistry. AIRE expression in TCs was supported by the RNA‐seq data in the TCGA‐THYM database. Furthermore, our bioinformatics approach to the recent single‐cell RNA‐seq data on human thymi has revealed that TCs hold molecular characteristics of multiple mTEC subpopulations. In contrast, TCs lacked the gene signatures for cTECs. We propose that TCs are tumors derived from mTECs. Thymic epithelial tumors originating from thymic epithelial cells (TECs) are classified into six types of thymomas  and thymic carcinoma (TC). Despite the clear distinction between cortical (cTECs) and medullary TECs (mTECs) in physiology, the cell of origin of each TET has remained enigmatic. In the current study, we confirmed the expression of AIRE, an mTEC‐specific transcriptional regulator, in most TCs at both protein and mRNA levels. Furthermore, our bioinformatics approach has revealed that TCs hold molecular characteristics of multiple mTEC subpopulations, suggesting their cell of origin as mTECs. While w2.2Immunohistochemical analyses were performed using formalin‐fixed paraffin‐embedded specimens. Antigen retrieval was performed by autoclaving the samples in citrate buffer (pH 6.0) for 30 min. We used anti‐AIRE  and anti‐cytokeratin AE1/AE3  antibodies. Rabbit polyclonal anti‐human AIRE antibody was raised by immunizing the 13 amino acids residing between the PHD1 and PHD2 domains of human AIRE. Dako Autostainer Link 48 was used following the manufacturer's protocol. For immunofluorescence, we used secondary antibodies conjugated to Alexa Flour 594 (Invitrogen) or Alexa Flour 488 (Invitrogen).2.3edgeR for the trimmed mean of M values (TMM) normalization.Seurat.We utilized an RNA‐seq dataset from the TCGA Project archive (TCGA‐THYM). The TCGA‐THYM cohort was comprised of 108 thymomas  and nine TCs. We used R package 2.4p value less than 0.05 was considered significant.Statistical significances of gene expression in the TCGA dataset (TC vs. other thymomas) were computed by one‐way ANOVA with post hoc Tukey HSD. Statistical significances of each gene expression in the scRNA‐seq data were computed by the Wilcoxon rank sum test. 3We examined a total of 90 samples diagnosed as TET Table . We appl+ cells were scattered throughout the tumor tissues like normal thymus. The proportion of AIRE+ cells was variable among the samples ranging from ~1% up to ~30%. By immunofluorescence, AIRE in TCs showed authentic nuclear dot morphology  compared with the conventional type B thymomas  contained a few AIREn Figure . Type ABl Figure . OverallAIRE expression from TCs among TETs, and type B thymomas showed a lower but significant AIRE expression , AIRE+ mature mTECs (mTEChigh), and post‐AIRE mTECs We further extended our view using the recent data obtained by scRNA‐seq. scRNA‐seq has revealed the existence of multiple subpopulations for mTECs as exemplified by the mTECs reminiscent of tuft cells in the gastrointestinal tract.s Figure . We alsos Figure . In cont4+ mTEClow genes was not high in TCs. Instead, it was relatively high in type A thymoma, a subtype historically considered to reflect the characteristics of mTECs. We consider that TCs and type A thymomas are derived from different subpopulations and/or developmental stages of mTECs; the former is closely associated with mature mTECs, whereas the latter is with immature mTECs.Our present study combining immunohistochemical analysis of AIRE with a bioinformatics approach has provided a novel insight into the pathobiology of TETs, especially for TCs. One possible reason for the discrepancy between the previous studyOur findings have raised a new possibility to the important question: Why do we seldom see the development of autoimmunity in patients with TCs in contrast to thymomas? The absence of AIRE expression from most thymomas has been discussed with the development of MG.In conclusion, our study has demonstrated that TCs hold the characteristics of mTECs with AIRE expression, suggesting their cell of origin as mTECs. However, expression profiling of TCs at single‐cell resolution is yet to be approached because our study is based on bulk RNA‐seq analysis. How our findings revealing the immunopathology of TCs can be utilized for the novel therapeutic strategy for this devastating tumor also awaits further study.Minoru Matsumoto: Conceptualization ; formal analysis ; investigation ; writing – original draft . Takuya Ohmura: Data curation ; software ; visualization . Yuto Hanibuchi: Investigation; visualization. Mayuko Ichimura‐Shimizu: Validation . Yasuyo Saijo: Investigation. Hirohisa Ogawa: Supervision ; validation. Ryuichiro Miyazawa: Supervision . Junko Morimoto: Supervision . Koichi Tsuneyama: Supervision ; validation. Mitsuru Matsumoto: Resources ; writing – review and editing . Takeshi Oya: Conceptualization; validation; writing – review and editing .This work was supported in part by Tokushima University President.All the authors declare no conflict of interest.The present study was approved by the Ethics Committee of Tokushima University Hospital. They also waived the requirement to obtain written informed consent from patients, given the study's retrospective nature .Table S1. The results of the re‐definition of the TCGA‐THYM samplesClick here for additional data file.


The dissolution of ARGFs matrix in 4% and 10% NaOH solutions were distinctly delayed to 28 d, as a consequence of the introduction of the MnO2-based nanoparticle layer, compared with nontreated ARGF occurring at 3 d in 4% NaOH solution. For the optimally modified ARGFs, the mass loss was controlled to 1.76% and 2.91% after 90 d of corrosion in 4% and 10% NaOH solutions, and the retention of tensile strength was increased by approximately 25%. With respect to the increment in alkali-resistant performance, the modified ARGFs can be promising candidates for wide applications in alkaline cement-based products.Glass fibers are widely used in cement-based precast products, given the reinforcing requirements for toughness and strength. However, inferior alkali resistance hinders the effectiveness of glass fibers in reinforcing cement-based materials. In this paper, nanoparticle coatings were applied on the surface of alkali-resistant glass fiber (ARGF) as a protective layer via the in situ chemical reaction of oleic acid (OA) and potassium permanganate (PP). The morphology and constituents of the as-prepared ARGFs were examined using scanning electron microscopy (SEM) and obtaining X-ray photoelectron spectroscopy (XPS) measurements. Mass loss and strength retention were investigated to characterize alkali resistance of modified ARGFs. Results showed that ARGFs could be optimally coated by a layer of MnO Cement-based materials are widely used in building materials, such as precast panels and blocks for walls. Given the requirement for the toughness and strength of practical precast products in buildings, glass fibers are commonly employed to reinforce the cement-based materials ,2, sinceThe ARGFs can generally be divided into two categories based on the manufacturing methods of the fiber materials. One common category is manufactured from formulating chemical compositions of a glass matrix with the optimum content of a rare earth or metal element ,6. The o2 on the alkali resistance of basalt fibers in 1 mol/L NaOH solution. The incorporation of 0.25% TiO2 by the mass of a basalt rock can form insoluble titanium hydroxide in the basalt fibers, which reduced the mass loss of the TiO2-basalt fibers at a corrosion age of 3 h by up to 50%. Lee et al. [2CO3 by the ratio of 1:1. However, the utilization of rare earth or metal elements in manufacturing ARGFs led to a fairly high cost for ARGF-reinforced cement products, which hinders the wide use of such ARGFs in common cement-based materials.With respect to the formulation of glass compositions, the incorporation of rare earth or metal elements can obviously increase the resistance to alkali attack in fibers ,9, such e et al.  utilizede et al.  reportedThe surface modification of glass fibers is taken to be a rational approach which could improve their alkali resistance. Diverse physical approaches, such as surface coating, sizing, and etching, and chemical strategies, such as oxidation, grafting, and chemical precipitation, have been put forward. Sayyar et al.  develope2 solutions to form surface coating. An awareness of the inferiority in nanomodification arose concurrently. Peled et al. [Recent advances have also exhibited promise in nanomaterials in boosting the mechanical properties and bonding performance of fibers ,17,18. Sd et al.  found thd et al. . One of The objective of the study herein is thus to improve the alkali resistance of glass fibers by modifying the fiber surface with nano materials, for instance, cement-based materials, that are resistant to high alkaline environments. Oleic acid and potassium permanganate were employed to synthesize manganese-dioxide-based nanomaterials on the surface of common commercial ARGF to further protect the fibers from alkali attack. The raw commercial ARGFs were treated with variable mass ratios of OA to PP (OA:PP) and variable modification times. The synthetic chemical compounds on ARGF after surface modification (ARGF-Mn) were analyzed using X-ray photoelectron spectroscopy (XPS) and obtaining scanning electron microscope (SEM) measurements. The alkali resistance performance of the ARGF-Mn was characterized by investigating the mass loss ratio and strength retention of the fiber materials in corrosive environments, prepared as 4% and 10% NaOH solutions. The outcomes of this study can provide new paths for the development of glass fibers with high alkali resistance.3 were employed as control fibers. Ordinary Portland cement P·O 42.5  corresponding to European CEM I 42.5 and standard quartz sands were used to measure the strength retention of glass fibers.Sodium hydroxide  and potassium permanganate  of analytical grade were used to synthesize nanomaterials on the surface of glass fiber. Commercial common ARGF AR 5325  of 1200 tex and 2.60 g/cm4 solution in a beaker. The OA:PP was set as 2, 6, 10, and 14, and the modification time was conducted as 12, 18, 24, and 30 h, respectively. Subsequently, the modified ARGF-Mn was washed with ethanol and distilled water, respectively, to remove unreacted residues, until the mixed solution became neutral, and was finally placed in a drying oven at 60 °C for 48 h.The ARGFs with 8–10 g were mixed with oleic acid and 0.005 g/mL KMnOARGF and ARGF-Mn of 12 pieces each (1 ± 0.0005 g) were placed in a vessel and then mixed with 4% and 10% NaOH solutions, respectively. After corrosion for 3, 7, 28, and 90 d, they were cleaned with distilled water until the pH value of the solution reached neutral. The cleaned glass fibers were immediately placed in a drying oven and dried at 60 °C to constant weight. The mass loss ratio of the glass fibers was determined using Equation (1), as follows:3), and Strength retention test complied with European standard EN 14649:2005 (E) . Mixing The surface morphologies of ARGF and ARGF-Mn, as well as the morphology evolution of these fibers in alkali attack were characterized by scanning electron microscope . Sputter coating of Au was conducted to enhance the conductivity of the samples. X-ray photoelectron spectrometer  was used for surface composition analysis. Full scan together with high-resolution scan spectra were measured. Mn2p was analyzed by XPSPEAK software (version 4.1) with Gaussian-Lorentzian function. Peak fitting was employed based on subtraction of Shirley background. The optimal fit was finally gained by comprehensive effect of peak position, full width at half maximum, and peak intensity.The surface property of ARGF-Mn could also be affected by the duration of surface modification. Given the optimum ratio of OA:PP, the effect of modification time on morphology of ARGF-Mn surface was shown in XPS measurements were conducted to determine the chemical substances formed on the surface of alkali-resistant glass fibers after modification. x+) for ARGF-Mn. Characteristic peaks of Mn2p3/2 and Mn2p1/2 elements were confirmed with respect to the binding energy located at around 642 eV and 653 eV, respectively. Such XPS characteristic peaks indicated the existence of chemical compounds containing Mn4+ and Mn2+ ions on the surface of ARGF-Mn. With respect to the Mn2p3/2 peak, it is shown that the binding energy of Mn4+ was 642.4 eV and that of Mn2+ differed in 642.2 eV and 641.0 eV [x+ ions with binding energies of 642.4 eV, 642.2 eV, and 641 eV corresponded to MnO2, MnSiO3, and MnO, respectively. Hence, chemical products of MnO2, MnSiO3, and MnO were generated on ARGF-Mn. Further fitting of the Mn2p1/2 peak clearly showed the presence of the MnO2 product on the surface of the modified fibers. This can be ascribed to the reduction role of KMnO4 as strongly oxidant in its reaction with oleic acid, as follows:641.0 eV . The che7+ from KMnO4 could be reduced to Mn4+, Mn3+, and even Mn2+ in the reaction between KMnO4 and oleic acid. The Mn4+ could precipitate in the stable form of MnO2, while the Mn3+ ions would undergo a dismutase reaction, by which they would be further reduced to Mn2+ and then oxidized into MnO2. The chemical reactions can be described by Equations (3) and (4) [The ions of Mn and (4)  accordin3+ ions would not be formed on the surface of ARGF-Mn, whilst those composed of Mn2+ could be generated. This was consistent with the fitting results in 2+ ions could exist in the form of MnO and MnSiO3. The presence of MnSiO3 substance can be attributed to the bonding of Mn2+ onto the surface of alkali-resistant glass fibers, in terms of -Mn-O-Si-, in which the element of silicon was provided by the glass fibers.Based on Equations (3) and (4), chemical compounds containing Mn4+ and Mn2+ was approximately determined by calculating the peak area of their XPS fitting curves in 2, MnO, and MnSiO3 were then estimated as 70.58%, 17.81%, and 11.61%, respectively, given the molar mass of these chemical compounds. Hence, the major synthetic substance on ARGF-Mn was MnO2. Such synthetic chemical compounds could greatly affect the alkali-resistant properties of glass fibers that provide physical protection for the fiber surface.The atomic weight of Mn2-based chemical compounds adjusting the alkali-resistant properties of glass fibers, the alkali-resistant performance of glass fibers was studied by measuring the mass loss and strength retention of the fibers corroded in NaOH solution for variable durations of degradation.With synthetic MnO−, silicate skeletons of glass fibers could be sensitively attacked by alkaline species, which led to the incontrovertible disconnection of silicate network. The connectivity of Si-O-Si was sharply diminished along with the increment of harsh alkaline ions transforming bridging oxygens into non-bridging ones, which accordingly accounted for the gradual strength corruption of glass fibers. Hence, the development of mass loss with time was obviously accelerated by enhanced NaOH concentration from 4% to 10% since it facilitated the alkali-silica reaction. For example, the mass loss ratio of ARGF corroded by 4% and 10% NaOH for 90 d was 2.51% and 4.97%, respectively. The corresponding chemical reaction could be expressed by Equation (5), as follows [ follows :(5)≡Si–OThe alkali-resistant performance of the glass fibers could be obviously improved by surface modification with an OA:PP ratio of 10 for 24 h. The mass loss ratio of ARGF-Mn corroded in 4% and 10% NaOH solution developed much slower with prolonged time than that of ARGF. The mass loss ratio of ARGF-Mn after 90 d of corrosion with 4% and 10% NaOH solution was dropped to 1.76% and 2.91%, respectively.2-based nanoparticles formed on the surface of ARGF-Mn, which can provide physical protection for the modified fibers from the NaOH-silica reaction. The protective nanoparticles decreased gradually during the corrosion from 3 d to 28 d by 4% NaOH solution and diminished at 90 d.2-based nanoparticles obviously accelerated with the increasing NaOH concentration from 4% to 10%. Substantial MnO2-based nanoparticles were dissolved at 3 d of corrosion in 10% NaOH solution, as shown in 2-based nanoparticles were peeled off from the surface of ARGF-Mn after 28 d of corrosion with 10% NaOH solution. For ARGF-Mn corroded in 4% NaOH solution, only a small portion of the protective layer was destroyed at 28 d, as shown in 2-based nanoparticles formed on the surface of ARGF-Mn were fully degraded after 90 d of corrosion in 4% and 10% NaOH solution. Still, the surface of ARGF-Mn at 90 d was less rough than that of ARGF, resulting from the enhancement of alkali resistance of ARGF-Mn due to the protection of MnO2-based nanoparticle layer.The degradation process of MnO2 based coatings on the surface of ARGF-Mn. Such a coating could resist corrosion, in alkali environments, from the hydration of cement particles and thus delay the corrosion of alkalis to glass fiber matrices. As a result, the strength of ARGF-Mn could be retained to a great extent before the nano-MnO2 was exhausted thoroughly. By contrast, ARGF was completely exposed to harsh alkali environments and suffered from aggressive alkaline attack. Thus, the Si-O network of ARGF was more vulnerable to be broken, leading to the rapid degradation of the strength of ARGF.2 were successfully constructed on the surface of ARGF by employing oleic acid and potassium permanganate as reactants, in order to further defend against alkali attacks on ARGF in alkaline environments. The mass loss ratio and strength retention of the modified ARGF-Mn exposed to NaOH solution were investigated for evaluating the effect of synthetic nanoparticles on the alkali resistance of the fibers. Based on the aforementioned, the following conclusions can be drawn:In this study, nanoparticles mainly consisting of MnO2 accounted for more than 70% of the nanoparticles, while MnO and MnSiO3 took up approximately 18% and 12%, respectively.The surface of ARGF was optimally coated with nanoparticles when modified with an OA:PP ratio of 10 for 24 h, with a broad and uniform distribution of the nanoparticles on ARGF-Mn surface successfully acquired. The synthetic compound of MnO2-based nanoparticles delayed the obvious dissolution of the ARGF matrix in 4% and 10% NaOH solutions to the corrosion age of 28 d, while nonmodified ARGF was obviously pitted at only 3 d of corrosion in 4% NaOH solution. After 90 d of corrosion in 4% and 10% NaOH solutions, the mass loss of the optimal ARGF-Mn was significantly reduced to 1.76% and 2.91%, compared with that of ARGF of 2.51% and 4.97%, respectively.The coating layer of MnO2-based coating layer established could effectively resist attacks of alkalinity from the hydration of cement particles on an ARGF matrix.With the optimum modification of the fiber surface, the retention of tensile strength of ARGF-Mn was approximately 25% higher than that of ARGF, illuminating the fact that the MnO


To describe the audiological characteristics and the type of intervention chosen on unilateral hearing loss cases in children and adolescents as well as to analyze correlations between the degree of hearing loss, the indication and the use of electronic devices.Observational, descriptive and retrospective study, carried out with information of 34 medical records from children and adolescents with unilateral hearing loss, assessed by two auditory rehabilitation services of medium complexity, throughout 2016 to 2019. Descriptive and Inferential statistical analysis were performed with the data.A predominance of profound sensorineural unilateral hearing loss in the right ear, of pre-lingual character, with 20.6% of malformations. The most adopted intervention was the hearing aid indication, although its use is low, regardless of the degree of the hearing loss. An association was found between the degree of the hearing loss and the healthcare professionals in indicating the use of the devices.The indication of hearing aids is the most frequent and this decision is influenced by the degree of the hearing loss, in which the devices are mostly indicated for mild to severe losses, with bigger divergence of conduct for profound hearing losses. Most cases were considered not suitable for Hearing Aid (HA) adaptation, being the management of classroom position the only care implemented.In the past, children with Unilateral Hearing Loss (UHL) were thought to normally develop in a way that such deficit was not seen as clinically relevant, school difficulties,5, limitations in communicative activities, especially in noisy environments, and difficulty in sound localization,7 have been shown in children with UHL, all of which may potentially harm language development. The implications of UHL can even go beyond those related to hearing- and language- disorders, interfering with the quality of life or even the intelligence scores of these children.New research has evidenced that among the consequences of UHL in the infant population, risk to language development has been highlighted as speech problems. In addition, in UHL due to malformations, the treatment possibilities include besides conventional HA by airway or bone conduction, bone anchored hearing aids or surgical reconstruction of the ear.Nevertheless, one cannot state that the use of HA in cases of UHL is unnecessary,11, have also been reported by literature as options to adapt UHL cases. The latter, however, does not present proven scientific evidence of its benefits in childhood.Other electronic devices, such as the Contralateral Routing of Signal (CROS), the Modulated Frequency system (FM system) and the Cochlear Implant (CI),3-5 . Thus, the analysis of how the auditory rehabilitation services are managing the cases of children with UHL is a relevant topic considering the plurality of conducts observed in the scientific literature.Recently, remarkable attention has been drawn to UHL rehabilitation due to factors such as the ever since decreased age of diagnosis as well as the results of studies highlighting the impacts of UHL on children's language and learning since 2004, of which includes the granting of HA and speech therapy. As for UHL cases in particular, the suitability to adapt HA, according to the instructions provided, is controversial and requires a justification so that the HA can indeed be concessed, e.g. difficulty in social integration. The aim of the therapeutic intervention in these cases is to stimulate auditory and linguistic abilities centered on family participation, relying on specific therapeutic strategies depending on the individual demands of each case,15.In Brazil, the rehabilitation of UHL is envisaged by the National Policy to Hearing Health Attention, a measure adopted by the Sistema Único de Saúde (SUS)In view of the above, the aim of this study was to describe the audiological characteristics and the type of intervention chosen in the cases of children and adolescent with UHL attended by auditory rehabilitation public services, as well as to analyze the correlations between the degree of hearing loss, the indication and the use of HA.This is an observational, descriptive and retrospective study, carried out with information from the medical records of two medium complexity hearing rehabilitation services of the SUS, both located in the state of Rio Grande do Sul.137 medical records of children and adolescents were analyzed throughout january 2016 to november 2019. Of these, 34  were found to have UHL, composing the sample of the study. The age range was 11 months to 18 years, mean age of 8.7 years, being 18 males and 16 females. The records were excluded only if there was any lacking information or incomplete data.The analyzed variables were: type and degree of hearing loss, affected ear, age of diagnosis, age of onset for hearing loss, use of HA and effectiveness of its use, image exams and speech therapy.. To define the degree of hearing loss, the already established classifications of both services were used, which differed according to the age of the subject. For children older than seven years old, the classification proposed by Lloyd and Kaplan was used and for those younger, Northern and Downs was used. Both information were extracted from the most recent audiological assessment available in the subject’s record. Both classifications on the hearing losses’ degree were used as this study portrays an analysis of medical records, thus, the authors decided to maintain the classification in order to respect the decision-making of the rehabilitation services studied.The UHL cases were classified according to the type of hearing loss in one of the three categories: sensorineural, conductive or mixedThe information regarding the use of HA, the indication and the type were verified in the otorhinolaryngological record. To address the use effectiveness of the devices, a total of eight or more hours was considered as effective. The information on this matter was examined through analyzing the data logging of the software of the device or in the last update in the medical record.The age of onset of UHL  and the presence of image testing were considered according to the otorhinolaryngological record. Only two subjects had imaging tests and the others had clinically proven malformations  after medical consultation. It’s important to highlight that further malformations in the remaining subjects of the study could only be discarded with imaging testing (Magnetic Resonance Imaging and/or Computerized Tomography of Maistoid). However, such exams were not requested and/or were not available at the services that received these subjects.Regarding the speech therapy, information related to whether the participants were exposed or not to therapeutic services as well as the main goals of the assistance were sought in the same records. The relation between the speech therapy method and the auditory abilities was not analyzed in this study.After the data were collected, a series of descriptive and inferential statistical analyses were performed. The correlation between the degree of hearing loss and the hours of HA use was measured using Spearman's correlation. Lastly, Fisher's exact test was used to verify the association among the decision of indicating the use of electronic devices, the degree of hearing loss and the age of diagnosis. A 5% level of significance was considered to all statistical tests, which were performed in the software R v.2.15.3.This study was approved by the Ethics in Research Committee of the institution of origin under the number 14804714.2.0000.5346 and all the ethical precepts established by Resolution No. 466/2012 of the National Health Council (NHC) that regulates research involving human beings were respected. Those responsible for the patients, when accessing the rehabilitation services, signed the Term of Acceptance to the Service Norms as well as a written informed consent terms, which states that all the medical record data may be accessed by the professionals of the service in a confidential manner, and can therefore be used for research.Sensorineural UHL was the most frequent type observed in 22 (65%) of the subjects, followed by conductive hearing loss, evidenced in eight (23%) subjects and mixed UHL was observed in four (12%) subjects. From the 34 children and adolescents included, 23  had the right ear affected while 11  exhibited UHL in the left ear. In regard to the age of onset of UHL, a pre-lingual character was identified in 23  subjects and post-lingual in 11 .From the studied sample, seven (20.6%) subjects were found to have malformations involving the ear. The age of the subjects at the moment of the audiological diagnosis varied between one month to 17 years, mean of 5.2 ± 5.1 years (mean ± standard deviation). 12 (35.3%) cases were identified in the Newborn Hearing Screening (NHS) and for these, the diagnosis occurred in the average of 3.3 months.The distribution of the subjects in regards to the adopted intervention can be observed in It’s noteworthy that all subjects were under audiological monitoring to monitor a possible progression of the hearing loss and/or involvement of the contralateral ear, but in 10 cases this was the only decision made as a conduct.The use of HA was indicated for most of the participants. HAs were indicated for 15 subjects  while three (16.7%) received the indication of CROS.In regards to speech therapy, 11 (32.3%) children and adolescents performed this intervention weekly. Seven cases were found to use HA and also attend weekly speech therapy, of which had the specific aims to train auditory abilities, stimulate speech/language and sensibilize family to ensure adherence and raise awareness of the HA use as well as the therapeutic process. The therapeutic aims for the three children that did not use HA but were undergoing speech therapy was to stimulate the speech/language development as well as the development of the auditory abilities of the unaffected ear. No information on the therapeutic aims was found in the medical records of the subjects who used CROS and were also attending speech therapy. The reasons why the rest of the subjects did not attend speech therapy sessions varied, according to their medical records, from presenting satisfactory evolution in terms of auditory and/or linguistic abilities or they lived far away from the auditory rehabilitation services in which this study was carried out.The information regarding the average hours of use of the electronic devices  was found in only 14 medical records. The amount of hours of use varied between 0.3 hours to 11.8 hours per day, a mean of 4.5 ± 2.9 hours (mean ± standard deviation), for both HA and CROSS.The Spearman correlation did not evidence significant correlation between the degree of hearing loss and the hours of use  (p=0.327).Also, The Fisher’s exact test was employed to analyze the association between the decision of the healthcare professionals in indicating the use of electronic devices and the age of diagnosis, although no statistically significant difference was obtained for this analysis (p=0.140). The cut-off point of seven years was considered for this analysis.By analyzing the data of the present study, we found that children and adolescents present predominantly profound sensorineural UHL in the right ear. The most opted conducted was the indication of HA, although the use of such devices were found to be low. Moreover, the decision of the healthcare professionals to indicate such devices is apparently influenced by the degree of hearing loss.,4,9,19 have also observed higher prevalence of profound sensorineural hearing loss in the infant population with UHL. A study showed worse results in speech and language assessments of children with severe to profound UHL when compared to other degrees of losses. Literature points that when the auditory deficit is higher, the deprivation of sensory stimuli in the auditory pathways tends to be higher as well and consequently, areas such as speech and language end up more compromised due to these aspects. Therefore, the degree of the hearing loss appears to clearly influence the language development of the subjects with UHL.Sensorineural hearing loss was the most frequently observed type of UHL, followed by conductive and, lastly, mixed. Profound hearing loss was evidenced in 41.2% of the sample. Other studies,3,10,21. The same predominance was also evidenced for malformation cases. It's important to consider that, in UHL cases, a shorter activation of the contralateral auditory cortex occurs, therefore, when it comes to right UHL, the left auditory cortex tends to be more deprived and so a higher risk to speech recognition and auditory localization may be expected . A Brazilian research carried out in the state of São Paulo found that children with right UHL showed more school and linguistic complaints than those with left UHL.In regard to the affected ear, a predominance of the right ear was observed in the present study, just like in the consulted literature. In some cases, the use of HA is necessary as normal hearing is hardly obtained even after the surgical reconstruction.A higher occurrence of referral to high complexity otologic services was observed in the children with UHL due to malformations in the present study . Such re. It’s necessary to discard all the adaptation possibilities of airway HA and thus, the investigation of the anatomical conditions of the external ear becomes more relevant. In this sense, audiological assessment may not be sufficient in these cases, requiring further exams such as image testing. In Still on the topic of ear-related malformations, bone anchored hearing devices such as the Bone Anchored Hearing Aid (BAHA) may also be a good alternative.It's important to emphasize that external ear malformations are generally associated with middle ear malformations and that, in 15% to 20% of the cases, internal ear malformations coexist with those of external ear as well. The justification for such fact may be found in the embryonary development, as the external and middle ear present the same embryonic origin while the internal ear develops separately in a previous gestational period. Literature believes that UHL is diagnosed later than bilateral hearing losses as they may appear imperceptible to others, such as the parents, for a higher period of time. Attention must be drawn to the 12 cases identified by the NHS, of which the diagnosis occurred on average at 3.3 months. This result in particular shows an important decrease of the age of diagnosis recommended by the international guidelines.In this study, the identification of the hearing loss occurred mostly in the preschool period, considering the mean age of diagnosis was 5.2 years, which is in agreement with other Brazilian study. As observed in .A high clinical variability was found in regards to the adopted intervention when it comes to auditory rehabilitating patients with UHL and, therefore, one can infer that such cases represent a challenge to doctors and audiologists, which is more evidenced when the target-public is of children. Another negative aspect related to the use of CROS is the decreased sound localization ability, although a possible benefit of this device would be to improve speech perception in silence or in situations where noise and speech are presented at front.Although the CROS can be indicated for children with severe to profound UHL, according to the international guidelines, evidence of its benefit is still lacking in literature for this population. Such concern is due to the fact that the presence of noise in the contralateral ear in environments such as the school could interfere in the normal ear and thus negatively affect hearing for these children,4. The fact that the UHL can become bilateral emphasizes the importance to closely monitor this population, as about 40% of these children may present hearing loss progression, both for the affected ear and the normal-hearing ear as well.As for the intervention possibilities in the UHL cases in children, the option of submitting the subjects only to the audiological monitoring in the present study was found to be expressive . The aim,28 refers to the time of daily usage of the electronic devices, both for HA and CROSS, which was very low. The main barriers regarding the use of HA in younger populations with UHL are: disconfort, poor perceiving of its benefits and social stigma. The use of FM system and preferential seats in classrooms appears to intensify furthermore the social stigma around children and adolescents with UHL.Another result evidenced in this research as well as in further studies on the topic. The use of two classifications on the hearing loss degree, used in the studied services and, also, the heterogeneity of the sample could perhaps have compromised the analysis for this variable, thus we can consider these the limitations of this study.No correlation between the degree of the hearing loss and the amount of hours of use of the electronic devices was found in the studied sample. Thus, it's not possible to affirm that the degree of hearing loss influences whether the children would use more or less the HA or the CROS, as it has been previously referenced in literature,9,11. In fact, the Joint Committee on Infant Hearing, in its last update emphasizes the necessity to include UHL in future expansions of the criteria for CI indication. Also, due to the inexistence of protocols to verify and validate the HA, especifically for UHL in children, the healthcare professionals are unable to assess the real benefit of this particular device in the various degrees of UHL, and thus its indication gets compromised.As observed in . Also, partnerships with HA companies would be a good possibility for these services in a way that the devices such as the HA, the CROS and FM system could be loaned for household testing and thus assist the healthcare professionals in decision-making.In the future, it would also be necessary that the Brazilian Public Services attempt to unify the assessment methods so that the conducts can be defined more properly and at an earlier stageThe type and degree of UHL in children and adolescents is predominantly profound, sensorineural, pre-lingual and affects the right ear, with presence of malformations in 20.6% of the cases. The most adopted intervention was the use of HA, although the use of such devices were found to be low, regardless of the degree of hearing loss. The decision to indicate electronic devices is influenced by the degree of hearing loss, being most frequently indicated in cases of mild to severe UHL. The conduct of professionals in cases of profound UHL is divergent, which reflects the lack of guiding protocols to help decision making in the management of these cases.