Mortierella alpina, one of the most representative oleaginous fungi, is the only certificated source of dietary arachidonic acid-rich oil supplied in infant formula. This study was conducted to improve TG production in M. alpina by homologous overexpression of diacylglycerol acyltransferase (DGAT) and linseed oil (LSO) supplementation. Our results showed that the homologous overexpression of MaDGAT1B and MaDGAT2A strengthened TG biosynthesis and significantly increased the TG content compared to the wild-type by 12.24% and 14.63%, respectively. The supplementation with an LSO concentration of 0.5 g/L elevated the TG content to 83.74% and total lipid yield to 4.26 ± 0.38 g/L in the M. alpina-MaDGAT2A overexpression strain. Our findings provide an effective strategy for enhancing TG production and highlight the role of DGAT in TG biosynthesis in M. alpina.Triacylglycerol (TG) with high-value long-chain polyunsaturated fatty acids is beneficial to human health; consequently, there is an urgent need to broaden its sources due to the current growing demand.  Triacylglycerol (TG), a type of neutral lipid comprising three fatty acyl moieties, is the major reserve of fatty acids (FAs) and the main form of energy storage in eukaryotes . The FAsMortierella alpina, one of the most representative natural producers of high-value LC-PUFAs, has already been used to produce dietary arachidonic acid-rich oil as a supplement in infant formula [M. alpina includes palmitic acid (C16:0), stearic acid (C18:0), oleic acid , linoleic acid , γ-linoleic acid , dihomo-γ-linolenic acid , and a large amount of arachidonic acid . Another indispensable FA, eicosapentaenoic acid , is usually present in M. alpina in trace amounts below 12 °C [M. alpina is an outstanding source of TG molecules with LC-PUFAs.The natural sources of TG-carrying LC-PUFAs are plants, oil crop seeds, marine fish, and oleaginous microorganisms that produce lipids over 20% of their dry cell weight (DCW) . Microbi formula . The FA c acid C1:0, oleicDGAT) catalyzes the final step in TG biosynthesis; it transfers the fatty acyl moiety from fatty acyl-coenzyme A (acyl-CoA) to diacylglycerol (DG) and commits DG to be stored as TG rather than being diverted for phospholipid synthesis [DGAT is divided into four families: DGAT1, DGAT2, WS/DGAT, and DGAT3 (CytoDGAT). Among these, two integral membrane-bound enzymes, DGAT1 and DGAT2, are ubiquitously found in eukaryotes without homology [M. alpina, the functional characterization and substrate preference of DGAT1 and DGAT2A have been explored in S. cerevisiae. MaDGAT2A could restore TG biosynthesis in S. cerevisiae H1246, and MaDGAT2B has also been applied to alter the FA and lipid composition [Diacylglycerol acyltransferase , and phosphatidylethanolamine (PE), which account for more than 95% of its total lipid content [There are thousands of lipid molecules in  sterols . The maj content . Benefit content . Further content ,19,20.DGAT has been applied to increase TG content and to accumulate desired FAs in several species, such as yeast [DGAT is one of the most effective enzymes for enhancing lipid accumulation due to its key role in TG biosynthesis. Supplementation with linseed oil (LSO) is an inexpensive and effective strategy for increasing the yield of high-value LC-PUFAs in M. alpina [M. alpinaDGAT on TG biosynthesis in vivo and the application of LSO supplementation on TG accumulation have not yet been explored.In previous studies, the overexpression of as yeast ,22 and aas yeast ,24. DGAT. alpina . HoweverMaDGAT1A, MaDGAT1B, MaDGAT2A, and MaDGAT2B, were overexpressed, and the effects of MaDGAT1B and MaDGAT2A on lipid profiles were investigated. Furthermore, LSO was administered to the M. alpina-MaDGAT2A overexpression and wild-type strains during fermentation. Subsequently, the total biomass of the mycelium was measured, and its lipid composition was analyzed to explore the effects of LSO supplementation on lipid biosynthesis. Thus, in this study, we attempted to provide a new perspective for obtaining more TG molecules with high-value LC-PUFAs for production applications.In this study, four identified genes, M. alpina ATCC 32222 was purchased from American Type Culture Collection . The uracil-auxotrophic strain M. alpina CCFM 501 was constructed by homologous recombination to knock out the ura5 gene encoding orotate phosphoribosyl transferase , based on the wild-type strain M. alpina ATCC 32222. The binary expression vector pBIG2-ura5s-ITs was constructed in a previous study and conserved in the E. coli top 10 strain [Agrobacterium tumefaciens CCFM 834 was a transfer DNA (T-DNA) donor for A. tumefaciens-mediated transformation. The uracil-auxotrophic strain, M. alpina CCFM 501, was used as the transformation recipient to construct a recombinant strain. CCFM refers to the Culture Collection of Food Microorganisms of Jiangnan University, School of Food Science and Technology, Research Center of Food Biotechnology.0 strain . AgrobacDGAT1 and DGAT2 from different species  listed in the National Center for Biotechnology Information ) as queries, the coding sequences of M. alpina DGAT1 and DGAT2 were obtained by homologous alignment in the genome database of M. alpina ATCC 32222 [Using several identified cDNA sequences of CC 32222 .M. alpina using the Trizol method and reverse transcribed to obtain cDNA. The target gene fragment was amplified by PCR, using cDNA as a template and KOD high-fidelity enzyme as an extension enzyme. The fragment was then ligated downstream of the His550 promoter, which was modified from the promoter of H4.1 genes from histone H4 isolated from M. alpina, in the vector pBIG2-ura5s-ITs and transferred into Agrobacterium tumefaciens by electroporation .Total RNA was extracted from M. alpina was cultured in a glucose yeast extract (GY) agar slant ) at 4 °C for preservation and spore production. Spores were collected with normal saline, filtered through Miracloth , and centrifuged at 12,000× g for 25 min at 4 °C. M. alpina mycelium was transferred from the GY agar slant into 100 mL of broth medium ) in a 250 mL flask and shaken for 48 h at 200 r/min and 28 °C. The mycelium pellets in suspension were dispersed using basic ULTRA-TURRAX  and transferred into a new broth medium . This step was repeated for three generations, each for 48 h. The dispersed mycelial suspension of the last generation was inoculated in the broth medium (with 30 g/L glucose) in a fermentation culture for 168 h at 200 r/min and 28 °C.A. tumefaciens, which had been transferred into the corresponding vectors, was streaked onto yeast extract peptone (YEP) agar medium. The single colony was inoculated into YEP medium with 100 μg/mL kanamycin and 100 μg/mL rifampicin at 30 °C for 36 h away from light until the medium turned orange; it was then transferred into a minimal medium (MM) with 7.8 g/L 2-(N-Morpholino)ethanesulfonic acid (MES)  at 28 °C for 36 h until the OD660 reached 1.5. Subsequently, A. tumefaciens was diluted and cultured in an induction medium (IM) containing 100 μg/mL acetosyringone and 7.8 g/L MES at an initial OD660 of 0.2 away from light at 30 °C for 12 h until the OD660 reached 0.8.M. alpina strain CCFM501 were collected approximately 12 h before the transfer of A. tumefaciens to the IM, as described above. The well-grown A. tumefaciens in IM was serially diluted to an OD660 of 0.2, 0.4, and 0.6 and mixed with 107−8 spores/mL of uracil-auxotrophic M. alpina spore suspension. Next, the mixture was coated onto induction agar medium, which was covered with cellophane membranes and supplemented with 0.1 g/L uracil for co-cultivation away from light for 24 h at 16 °C and, then, for 24 h at 28 °C. Subsequently, the cellophane membranes were transferred to an uracil-free synthetic complete (SC) agar medium with 100 μg/mL cefotaxime and 100 μg/mL spectinomycin for screening after the appropriate density of mycelium and bacteria had grown on the membrane. Mycelia from the fungal colonies were picked and transferred to new uracil-free SC agar plates one by one. This step was repeated thrice to obtain genetically stable transformants.The spores of the uracil-auxotrophic M. alpina according to the manufacturer’s instructions. As previously described, the presence of T-DNA in the genome of transformants was identified using the universal primers Hispro F1 and TrpC R1. The PCR product was collected by gel extraction and purified using a DNA purification kit  for sequencing.A Biospin fungal genomic DNA extraction kit  was used to extract genomic DNA from v/v) from approximately 30 mg of powder, and methyl-esterified with 4% (v/v) H2SO4 in methanol for further analysis. The FA profiles and total lipid yield were analyzed in the form of FA methyl ester using gas chromatography-mass spectrometry (GC-MS) analysis  with a DB-WAXetr column . Pentadecanoic acid (C15:0) was used as the internal standard for relative quantification, and the temperature program was performed as previously described [After 168 h of fermentation, the fresh mycelia were obtained by fast filtration using a paper filter, washed with distilled water to remove the culture medium, and frozen at −80 °C. Then, they were dried in a vacuum freeze-dryer overnight to quantify the DCW. The freeze-dried biomass was ground into a fine powder for total lipid extraction. All the lipid types were hydrolyzed with 4 mol/L hydrochloric acid solution, extracted with methanol:chloroform  for further analysis.To reduce contamination of the residual cell contents, a modified methyl tert-butyl ether (MTBE) extraction method was used for lipid extraction for untargeted lipidomics analysis ,28. BrieLipidomics analysis was performed using a Dionex UltiMate 3000 UPLC system  coupled with a Q Exactive Orbitrap mass spectrometer . The lipid extract was separated on a Waters Acquity UPLC BEH C18 column  maintained at 40 °C. We used a binary solvent system comprising mobile phases A (60:40 acetonitrile/water solution containing 10 mM ammonium formate and 0.1% formic acid) and B (90:10 isopropanol/acetonitrile containing 10 mM ammonium formate and 0.1% formic acid). The elution gradient was as follows: in the positive ion mode, 0.0–3.0 min, 0–70% B; 3.0–26.0 min, 70% B increasing linearly to 90%; 26.0–27.0 min, 90% B; 27.0–28.0 min, 90% B decreasing linearly to 0%; and 28.0–30.0 min, 0% B; in the negative ion mode, 0.0–3.0 min, 10–70% B; 3.0–12.0 min, 70% B increasing linearly to 80%; 12.0–15.0 min, 80–95% B; 15.0–15.1 min, 95% B decreasing linearly to 10%; and 15.1–18.0 min, 10% B. The mobile phase flow rate was 0.25 mL/min, the autosampler temperature was 15 °C, and the injection volume was 2 μL.5, and a relative standard deviation (SD) of >30% were excluded. The ID quality filters were selected as grades A and B. The adduct ions were +H and +NH4 in the positive ion mode and −H, +HCOO, and −2H in the negative ion mode. The content of a particular lipid class is obtained by dividing the total peak area of that lipid class by the total peak area of all lipid molecules.The MS1 and MS2 data were acquired using a Q-Exactive Orbitrap mass spectrometer through data-dependent acquisition in the range of 200–1200 for both the positive and negative ion modes. The detailed parameters were set as previously described, including lipid identification and alignment . Lipid ip < 0.05.All data are presented as means ± SD, representing at least three independent experiments. SPSS software version 20.0  and GraphPad Prism version 8  were used to perform statistical analysis and visualization. Paired two-tailed, two-tests were used for two-group comparisons, and one-way analysis of variance with Tukey’s test was used for multiple comparisons. Statistical significance was defined as M. alpina DGAT1A, DGAT1B, DGAT2A, and DGAT2B genes were obtained from the genome of M. alpina ATCC 32222 based on the results of the sequence alignment and ligated to the binary expression vector pBIG2-ura5s-ITs. The T-DNA regions containing MaDGAT1A, MaDGAT1B, MaDGAT2A, and MaDGAT2B, with the selection marker urs5s, were inserted into the genome of M. alpina CCFM 501 by A. tumefaciens-mediated transformation. Several transformants with genetic stability were successfully obtained, and segments of MaDGAT1A, MaDGAT1B MaDGAT2A, and MaDGAT2B were amplified from the genome of the transformants by PCR. M. alpina-MaDGAT2A-3/4/6/8.The M. alpina-MaDGAT1A recombinant strains, eleven M. alpina-MaDGAT1B recombinant strains, six M. alpina-MaDGAT2A recombinant strains, and three M. alpina-MaDGAT2B recombinant strains. The complementary-phenotype strain obtained by transforming the vector pBIG2-ura5s-ITs into the uracil-auxotrophic strain was used as the negative control  for 7 d, we separated and identified the lipids of M. alpina-MaDGAT1B, M. alpina-MaDGAT2A, and M. alpina ATCC 32222 by UPLC-QE-MS/MS, and the repeatability and reliability of the data were confirmed by principal component analysis , increased by 12.24% and 14.63%, respectively. The DG content in M. alpina-MaDGAT1B/2A decreased from 10.71% to 3.70% and 3.08%, respectively, compared with that in M. alpina ATCC 32222  to 11.28% and 10.33%, and the PE content decreased from 3.37% to 2.62% and 2.37%, respectively , TG (20:4_20:4_20:4), TG (20:3_20:4_20:4), and TG (18:3_20:4_20:4); however, the content of these TG molecules were reduced in M. alpina-MaDGAT1B/2A compared to wild-type strain. In contrast, TG (16:0_20:3_20:4), TG (16:0_18:1_18:3), and TG (18:0_18:0_20:4) in M. alpina-MaDGAT1B and TG (18:1_18:1_18:3), TG (18:1_18:1_18:2), and TG (16:0_18:1_18:1) in M. alpina-MaDGAT2A exceeded their content in the wild-type strain. The species of major DG and PC molecules were much less than those of the TG molecules  and DG (18:3_18:2). A similar situation was observed with the PC molecules, where the percentages of PC (20:4_20:4) and PC (16:0_20:4) were higher in the wild-type than those in M. alpina-MaDGAT1B/2A, but the percentages of PC (18:1_18:2) and PC (16:0_18:1) were higher in M. alpina-MaDGAT1B/2A than those in the wild-type  until the concentration reached 0.5 g/L.M. alpina-MaDGAT2A reached 4.26 ± 0.38 g/L. Interestingly, the increase in total biomass and total lipid yield in M. alpina-MaDGAT2A were greater than those in the wild-type strain. Moreover, the TG content reached 83.74% in M. alpina-MaDGAT2A with 0.5 g/L LSO supplementation; however, the TG content of the wild-type strain showed a slight decline from 71.93% to 71.53%  supplementation in recombinant yeast [In a previous study, we performed a lipid analysis of analyses ; howeverprofiles . These rnt yeast .M. alpina-MaDGAT1B and M. alpina-MaDGAT2A with that of wild-type demonstrated that MaDGAT1B and MaDGAT2A could significantly promote the conversion of DG to TG but not the synthesis of phospholipids, where the content of PE and PC decreased (MaDGAT1B and MaDGAT2A enhanced the total lipid yield, regardless of a slight decrease in biomass, compared to the wild-type strain (M. alpina-MaDGAT2A, indicating that MaDGAT2A provides more power to produce TG and that MaDGATs may play different roles in TG biosynthesis. The different substrate preferences of MaDGATs lead to their different effects on TG synthesis. NoDGAT1A knockdown and overexpression appreciably impacted TG accumulation in N. oceanica; hence, the overexpression of NoDGAT1A increased TG yield by 47% [NoDGAT2s that possessed distinct substrate specificity were used to produce TG molecules with tailored LC-PUFAs [PtDGAT1 is superior to the other five PtDGATs in biosynthesizing TG [MaDGAT overexpression is a highly effective metabolic engineering strategy to increase both total lipid and TG production.Furthermore, comparing the TG content of ecreased . Anothere strain . It is wd by 47% . On the LC-PUFAs . The resizing TG . In summM. alpina-MaDGAT2A was greater than that of the wild-type (M. alpina-MaDGAT2A, and the increase in DG content in the wild-type strain was greater than that in M. alpina-MaDGAT2A (MaDGAT2A can enhance lipid production and TG biosynthesis with LSO supplementation more than that in the absence of LSO, but the wild-type strain may have difficulty converting such a lot of LSO. Peony seed oil was supplied to M. alpina in previous study, and the EPA production was improved to 588.5 ± 29.6 mg/L in a 5-L bioreactor with a Micromonas pusilla delta-6 desaturase gene overexpressed, which resulted in a 26.2-fold increase compared to EPA production in wild-type M. alpina [MaDGAT2A overexpression strain did not obtain such a great boost. A possible explanation for this result may be that the high intrinsic TG content restricts enhancement range.After supplementation with LSO, the increase in the total lipid yield of ild-type . The TG MaDGAT2A . HoweverMaDGAT2A . These f. alpina . NeverthM. alpinaDGATs in enhancing TG production and lipid accumulation. With LSO supplementation, TG production by M. alpina-MaDGAT2A could be further enhanced by increasing substrate availability. Our investigation offers new strategies for improving TG production from the perspective of metabolic engineering and substrate supplementation while also supporting industrial TG production with LC-PUFAs. In future work, rewriting the carbon flux into FA synthesis to provide sufficient substrate for TG synthesis and blocking TG breakdown pathways by multi-gene operation system are potential strategies to further increase LC-PUFA and TG production. Furthermore, differences in the substrate preference of MaDGATs could be exploited to biosynthesize TG molecules with tailored LC-PUFAs.Overall, our findings revealed the potential and high application value of


The transient evoked otoacoustic emissions (TEOAEs) have been widely used in neonatal hearing screening.To compare the TEOAEs in newborns at term and preterm vis-à-vis the following variables: ear side, gender, frequency spectrum and gestational age.th day of life (41 newborns at term and 25 premature babies), we recorded TEOAEs. All the individuals did not have risk indicators for hearing loss.By means of a cross-sectional cohort of 66 newborns up to the 28There was a signal/noise ratio improvement with frequency increase. No differences were observed between genders and between the ears, but there were differences among the children born at term and preterm in the frequency bands at 3 kHz and 4 kHz.The TEOAEs test is important for assessing the peripheral auditory system of newborns at term and preterm, making it possible to have responses regardless of gender and gestational age. Auditory system integrity is important for an individual's communication and social interaction. Hearing enables the child to develop the verbal mode of communication, so it is essential to screen hearing at birth.Many techniques are used for the evaluation of hearing sensitivity and, among them, we have otoacoustic emissions. There is an acoustic energy that can be measured in the external ear canal, spontaneously or evoked by sound stimuli. When evoked, these emissions are usually classified according to the generating stimulus: transient, frequency stimulus and distortion productIn his first publication on the subject, KempIn recent years, the recording of transient evoked otoacoustic emissions has been widely used in neonatal hearing screening of newborns without hearing risks, using low intensity stimuli, with a wide range of frequencies and in a short period of timeChapchapIn a study assessing the peripheral hearing of 157 full term and preterm newborns, normal and small for gestational age, using transient otoacoustic emissions, Garcia et al.According to the organizations involved with pediatric hearingIn normative data measurement for transient otoacoustic emissions, some authors have observed a decrease in otoacoustic emissions with increasing ageth week of gestational age and that after that age, the record becomes similar between full term and preterm newborns.Tognola et al.In a recent studyDuarte et al.Given the variety of findings found in the comparison of full term and preterm newborns, we felt the need to study transient otoacoustic emissions in full term and preterm newborns, without any hearing risk, in order to try to understand cochlear function maturation in this age group that has no risk of hearing.To compare the signal-to-noise ratio of transient evoked otoacoustic emissions in full term and preterm newborns, vis-à-vis the following variables: ear side, gender, frequency spectrum and gestational age.The present investigation is a cross-sectional cohort study, developed after approval by the Ethics in Research Committee, under protocol 11447/2007.To do this study, an evaluation was scheduled within the first 28 days of life, for the analysis of transient evoked otoacoustic emissions (TEOAE) of 66 infants in good health. All newborns were born and recruited at the University Hospital of the Medical School of Ribeirão Preto - University of São Paulo, and the test was scheduled at the time of hospital discharge in the child's release card. The mothers were informed about the study and all signed an informed consent form.Two groups were formed: Group 1, with 41 healthy and full term newborns, 29 females and 12 males, with gestational age of 37 to 42 weeks; and Group 2, with 25 healthy preterm newborns, 13 females and 12 males, between 32-36 weeks of gestational age.•Family history of permanent hearing loss in childhood;•NICU stay for more than five days, or the presence of any of the following risk factors, regardless of length of stay: mechanical ventilation, exposure to ototoxic drugs and hyperbilirubinemia serum level requiring exchange transfusion;•Congenital infections such as cytomegalovirus, herpes, rubella, syphilis, and toxoplasmosis;•Craniofacial anomalies, including disorders in the ear canal, ear marks, ear pits, and temporal bone abnormalities;•Physical findings associated with other syndromes related to sensorineural hearing loss or permanent conductive hearing loss;•Syndromes associated with progressive or late-onset hearing loss, such as neurofibromatosis, osteopetrosis, Usher syndrome, and other frequently identified syndromes including Waardenburg, Alport, Pendred, Jervell and Lange - Nielson;•Neurodegenerative disorders, such as Hunter syndrome, or sensorimotor neuropathies, such as Friedreich's ataxia and Charcot-Marie-Tooth syndrome;•Postnatal infections associated with sensorineural hearing loss, including bacterial and viral meningitis;•Traumatic brain injury, especially temporal bone fractures that require hospitalization;•Children who failed the newborn hearing screening performed by TEOAE.All children selected for this study had no risk indicators for hearing lossThe TEOAE was performed in a soundproof booth with the baby under natural sleep or quiet.The equipment used was the Intelligent Hearing System SMART-EP. Newborns were evaluated in both ears. The stimulus was a click at the intensity of 80 dB SPL and at least 500 stimulations were performed on each test. Response reproducibility observed was of at least 50% correction between the A and B curves, and was regarded as passes in the newborn hearing screening, the presence of emissions by at least three frequency bands, considering the signal-to-noise ratio (S/N) greater than or equal to 3 dB at 1 kHz and 1.5 kHz and greater than or equal to 6 dB at 2 kHz, 3 kHz and 4 kHz.t-test with the GraphPad Instat software.The data was analyzed by the Student's The signal-to-noise ratio of otoacoustic emissions was described as average, minimum, maximum and standard deviation.To compare the signal-to-noise ratio of otoacoustic emissions between the right and left ears of the individual and between genders and ages, we employed the variance analysis (ANOVA), considering that the data followed a normal distribution.p < 0.05.The significance level was 95% with We observed the presence of TEOAE in all newborns evaluated.The averages of the signal-to-noise ratios (dB SPL) for the TEOAE of the preterm newborns, for different frequencies were: 4.39 (1 kHz); 10.39 (1.5 kHz); 11.77 (2 kHz); 12.54 (3 kHz) and 9.54 (4 kHz); and for full term newborns they were: 3.49 (1 kHz); 10.28 (1.5 kHz); 11.58 (2 kHz); 15.1 (3 kHz), and 12.65 (4 kHz).For both groups, one can observe an increase of the amplitude response with increasing the frequency tested, except at 4 kHz.In In the comparison between the ears of full term newborns we found a similar signal-to-noise ratio mean value in most frequencies, without statistical significance in the frequencies of 1 kHz; 1.5 kHz; 2 kHz and 4 kHz. There was only statistically significant difference in the 3 kHz frequency, with a better left-side mean value when compared to the right side .In comparing the ears of preterm newborns we found a similar signal-to-noise ratio mean value for the TEOAE test in all frequencies analyzed, without statistical significance .In comparing the signal-to-noise ratio of full term and preterm newborns, we found similar mean values in the frequencies of 1 kHz; 1.5 kHz and 2 kHz, with no statistically significant difference. There was a statistically significant difference in the frequencies of 3 kHz and 4 kHz, with a better mean value for the full term group than among the preterms .In comparing the signal-to-noise ratio of full term newborns, we found a statistically significant difference for the frequencies of 1.5 kHz and 2 kHz, with better mean values for females than for males in most frequencies analyzed .In comparing the signal-to-noise ratio of preterm newborns, we found a statistically significant difference for one frequency band only - 1.5 kHz, with better mean values for males than for females, with similar values in the remaining frequencies .When we studied the TEOAE signal-to-noise ratio (in dB SPL) in full term and preterm newborns of both genders, there was statistical significance only at 1 kHz and similar amplitudes in the remaining frequencies .The results showed the presence of TEOAE in 100% of full term and preterm infants. These results confirmed the findings by PeckThe use of this test for hearing screening in newborns was effective, confirming reports from previous studiesIn a study of cochlear function in neonatal ICU babiesThe present study also confirmed major differences in the higher frequencies, wherein maturation occurs later - as observed in another paperA similar study was carried outIn this study, we observed that, in comparing the ears, full term and preterm subjects had similar responses in both ears, and statistical significance was found only in the frequency of 3,000 Hz in the full term group, with better values on the left side. We cannot infer a better response from any of the ears because the findings were very similar. In comparing genders, the females of the full term group had better mean values, with statistically significant differences for the frequencies of 1,500 Hz and 2,000 Hz. The preterm group had a statistically significant difference at the 1,500 Hz frequency only, with males having better amplitudes. When analyzing the full term and preterm groups, we found no differences between genders for any frequency. The same was not reported by other authorsAn evaluation of 582 full term neonates by means of otoacoustic emissions 48 hours after birthThese findings enable us to confirm that the TEOAE test provides important information on the peripheral auditory system of newborns at full term and preterm, and also because of the relative ease of recording it should be a method of choice in newborn hearing screening protocols for cochlear acuity assessment.The examination can be performed easily in newborns irrespective of gestational age.From the result analysis we conclude that in comparing genders and ears, there was no difference in the signal-noise ratio between genders and the right and left ears vis-à-vis the TEOAE test in the groups of newborns at full term and preterm.Improvements were seen in the signal-noise ratio with increasing the frequency bands evaluated.With respect to gestational age, there were significant differences between the full term and preterm groups in the frequency bands of 3 and 4 kHz, with better responses for full term newborns.


To investigate the behavior of silicone oil (SiO) at a steady equilibrium and during saccades in pseudophakic highly myopic eyes with posterior staphyloma with and without an encircling band and compare it to behavior in emmetropic eyes. The SiO–retina contact area and shear stress were calculated by computational fluid dynamics.A numerical model of an emmetropic eye and a myopic eye with and without scleral band underwent a saccade of 50°/0.137 s. The vitreous chamber surface was divided into superior and inferior 180° sectors: lens, pre-equator, post-equator, and macula. SiO–retina contact was evaluated as a function of fill percentages between 80% and 90% for standing, 45° upward tilt, and supine patients. Maximum and average shear stress were calculated.Overall, SiO–retina contact ranged between 40% and 83%; fill percentage varied between 80% and 95%. Neither the encircling scleral band nor the staphyloma significantly affected the SiO–retina contact area, although the presence of a scleral band proved disadvantageous when gazing 45° upward. The inferior retina–SiO contact remained below 40% despite 95% SiO fill. The SS significantly increased at the scleral band indentation and decreased elsewhere. The staphyloma greatly reduced shear stress at the macula.The presence of a myopic staphyloma reduces shear stress at the macula but does not alter SiO–retina contact significantly. The apposition of a 360° scleral band may reduce SiO–retina contact at least in some postures and increases the SS at the indentation.Assessing SiO–retina contact when vitreous chamber geometry changes according to pathologic or iatrogenic modifications allows accurate prediction of real-life tamponade behavior and helps explain surgical outcomes. Pars plana vitrectomy involves removal of the vitreous gel and substituting it with gases or liquids, collectively defined “tamponades,” that are intended to contact the broadest possible extension of the retinal surface, especially for retinal detachment repair.Although all vitreous substitutes exert surface tension at the retinal interface, it is most likely the displacement of aqueous from retinal tears that prevents retinal re-detachment, whereas several other factors play important roles: surface tension, gravity, buoyancy, patient positioning, and vitreous chamber shape.Previous studiesThe present study used CFD to evaluate static and dynamic contact of SiO tamponade with the retinal surface of myopic eyes, in the presence of posterior staphyloma and an encircling band, and compared it to emmetropic geometry.–The numerical simulations were performed using the open-source library OpenFOAM, which has been successfully adopted in various contexts, including the fluid dynamics of the eye.−1), and thus is assumed to behave as a Newtonian fluid. Hence, our computational model adopted Newtonian constitutive laws for both SiO and aqueous.SiO (1000-mPa·s dynamic viscosity) was utilized as a tamponade. Because the SiO used here is composed of 100 % of polydimethylsiloxane, it has a negligible elastic component and a nearly constant shear viscosityThe multiphase flow was simulated, under the hypothesis of incompressible, isothermal, and immiscible fluids, by means of the volume of fluid technique (VOF), which is based on coupling the continuity and momentum equation with an additional conservation equation for the fluid mixture.t is time, θ is angular displacement, and the coefficients c1,…,c4 were chosen to reproduce a realistic saccade •Pre-equatorial retina, the retinal surface anterior to the geometric equator (red)•Post-equatorial retina, the retinal surface posterior to the geometric equator and between the equator and the macula (blue)•Macula, the retinal surface centered on the geometric posterior pole and extending for an angle of 20° anteriorly Each of above regions was further divided into superior and inferior halves.The main study outcome measures include retinal contact measured as the percentage value of retinal surface wet by SiO and the shear stress at the retinal surface.P < 0.05 was considered statistically significant.ANOVA was applied to statistically evaluate the significance of shear stress values at different locations, as well as to compare the results obtained for the various regions. SiO–retina contact is reported as a function of patient position and SiO fill percentage in The superior pre-equatorial retina a–3c showThe presence of an encircling band proved disadvantageous for a standing patient with 95% fill d. FigureA 90% SiO fill was considered representative of the “real-world” post-operative condition; interestingly, the presence of an encircling band reduced the fraction of total SiO–retina contact for a standing patient a, but itOverall, the inferior quadrants showed limited SiO contact for a standing patient, even with a 90% tamponade fill , and theThe macula completely contacted SiO only for a standing patient and was entirely wetted by aqueous at the 45° upward tilt and supine positions, regardless of the presence of an encircling band. Supine position almost completely prevented SiO from getting into touch with the post-equatorial retina, including the macula 45–6.In order to characterize the tangential forces exerted by the SiO on the retina during the saccade, we considered the case of a standing patient with a 90% fill and compared the three geometries, analyzing the values of shear stress reached during the simulation time. The maximum shear stress attained in each region during the entire observation time is plotted in The average shear stress over each region and during the entire observation time, displayed in Reaching and maintaining the broadest possible contact between the retina and SiO have been regarded as being pivotal in reducing the risk of retinal re-detachment. SiO effectively keeps aqueous away from retinal tears, preventing its leakage into the subretinal space, and reduces the volume of aqueous reservoirs where inflammatory mediators pool, thus decreasing the risk of proliferative vitreoretinopathy (PVR).The present paper describes SiO statics and dynamics and its interaction with the retina under the most frequent pathologic or iatrogenic conditions altering the vitreous chamber contour , compared to an idealized emmetropic eye. In our CFD model, SiO fill percentage proved to be the most important single factor impacting the extension of the retinal contact. Its increase from 80% to 95% in volume significantly increased the overall retinal contact area regardless of patient positioning and ocular geometry variations .Interestingly, an 80% SiO fill contacted only a little more than 50% of the entire retinal surface regardless of the eye geometry and positioning, and even a 95% volume fill left about 20% of the retinal surface untouched . It shouThe apposition of a scleral band in association with pars plana vitrectomy has been regarded as offering several benefits. It would improve the contact between SiO bubble and the retina,,Our model indeed shows that the presence of an encircling band does not increase the amount of retina in touch with the tamponade . TherefoWhen analyzing the retinal sectors separately, the inferior retina showed limited contact with SiO; up to a 95% fill rate and the presence of a scleral band significantly improved only pre-equatorial retinal contact when the patient was gazing upward b. As expOverall, our model does not support the hypothesis that 360° scleral indentation improves SiO–retinal contact, except for a few notable exceptions regarding specific sectors and eye positions of uncertain clinical relevance. More importantly, even when a 90% SiO fill was considered, none of the retinal sectors maintained a satisfying tamponade contact, regardless of the positioning.Myopic staphyloma did not impact SiO–retinal contact significantly in any of the cases considered. The macula maintained good contact with SiO only for a standing patient, whereas all of the other positions left it completely wetted by aqueous even with 90% SiO fill; the staphyloma and scleral band made no difference in any such cases . Shear sIn summary, our model shows that the highest possible SiO fill is mandatory, as even 90% fill leaves almost 30% of the retinal surface untouched, regardless of a patient's position. Moreover, the current outcomes suggest that the introduction of new tamponades contacting the entire retina would be highly desirable.


Gaps between performance and capability were observed for IADL but not for BADL. This study distinguishes between ADL performance and capability in individuals with mild dementia and is the first to identify the IADL-specific gap between these two components; pertinent targeted interventions are vital in closing this gap. Environmental adjustments are important to improve QOL of persons with mild dementia.Persons with mild dementia can effectively maintain and improve their quality of life (QOL) by regularly performing their daily activities. However, research on activities of daily living (ADL) in this population often fails to distinguish between ADL performance and ADL capability, that is, actual independence in daily lives and potential independence in an ideal setting. This study aimed to identify the potential gaps between ADL performance and capability in individuals with mild dementia. A total of 137 community-dwelling older adults (aged ≥ 65 years) who had been diagnosed with dementia and assessed as 0.5 and 1 on a global clinical dementia rating (CDR). Participants were evaluated for basic ADL (BADL) and instrumental ADL (IADL) using the Hyogo Activities of Daily Living Scale (HADLS). Around 35 individuals who met the inclusion criteria were enrolled in the study. BADL performance and BADL capacity were not significantly different ( Dementia has become a global challenge, with more than 55 million people worldwide currently suffering from dementia, a figure that is expected to increase to 139 million by 2050 [Dementia is a disease characterized by the chronic progression of core symptoms such as memory impairment and disorientation . Memory ADL consists of basic ADL (BADL), such as self-care, and instrumental ADL (IADL), for example, preparing meals, cleaning, laundry . In termDiscrepancies between ADL performance and ADL capacity, that is, between the actual performance of ADL in the elderly in everyday living environment and what they can achieve in a controlled environment with the necessary tools  and human factors such as appropriate support, relationships and attitudes, have long been noted in ADL assessments of the elderly . On the It is essential to distinguish between ADL performance and capacity as both can provide pertinent assessment information by demanding ADL-targeted interventions to help persons with dementia achieve full ADL potential in their daily routines ,13.This study’s objective was to determine whether a gap between ADL performance and ADL capacity exists among individuals with mild dementia to develop ADL intervention guidelines for this population.This cross-sectional study aimed to determine the status of ADL implementation in community-dwelling individuals with mild dementia.The authors invited more than 100-day care centers in four regions of Japan to participate by mail. 26-day care centers expressing interest in the study were visited by the authors to convey additional detailed information about the study. A day care center is a place where people living at home in the community have the opportunity to engage in activities and socialize during the day. The participants of this study are those who use those day care centers. In order to carefully assess the subject’s usual ADL performance, those who had been using the day care center for at least one month were included. Candidates were recommended by the day care center’s manager. A total of 137 community-dwelling older adults (age ≥ 65 years) who had been diagnosed with dementia using the long-term care insurance system of Japan were evaluated. They were enrolled if they had a global clinical dementia rating (CDR) of either 1 or 0.5 and were mobile indoors without the use of a wheelchair. Candidates were excluded if they lived alone, had a severe neurological or musculoskeletal disorder that interfered with physical activity, severe problems with cardiopulmonary or circulatory function, severe vision or hearing loss, or a history of aggressive or violent behavior noted by the care staff. When diagnosed with dementia, participants did not undergo formal cognitive screening, diagnostic imaging, or pathological testing to comprehensively explore their diagnosis and disease stage owing to personal financial constraints.The study subjects are people with dementia. Caregivers are in a position to evaluate them along with occupational therapists. Caregivers included in the study were those working at the same day care center as the participants, with no restriction on the caregivers’ profession, who directly took part in the care of the participants, and were most aware of information about the participants, including physical and psychological symptoms over the past month. Candidates were asked to provide written informed consent; when they were unable to provide it, their substitutes were asked to sign the consent form on their behalf.Participants were examined using the Hyogo Activities of Daily Living Scale (HADLS) . The HADCognitive function was evaluated using the Mini-Mental State Examination (MMSE) and was conducted through face-to-face interviews with patients . CDR evaData were analyzed using IBM SPSS Statistics 26.0  for Macintosh. All variables were confirmed for a non-normal distribution prior to analysis. The Wilcoxon signed-rank test is used to test for significant differences between two conditions of an independent variable data is collected from the same participants.p < 0.05 being considered statistically significant.In this study, as intra-individual changes due to environmental adjustments in ADLs, comparisons of median the performance and capacity scores for BADLs and median the performance and capacity scores for IADLs were analyzed using the Wilcoxon signed-rank test, with The study was conducted in accordance with the Declaration of Helsinki, and the protocol was approved by the Research Ethics Review Committee of the Health Science University (accession no. 1). The aims and methods of the study were explained to the participants and their substitutes. All participants gave their informed consent for inclusion before they participated in the study.Of the 137 patients, 35 individuals who met the criteria were enrolled . A total of 12 and 23 participants were rated as having CDR 0.5 and 1, respectively. The median (IQR) MMSE was 21.0 (19.0–22.0).On the HADLS, participants’ median BADL performance and BADL capacity were 4.5 (2.9–10.2) and 4.5 (2.8–10.0), respectively. In comparison, the median IADL performance and IADL capacity were 31.9 (24.9–37.5) and 25.3 (21.4–28.8), respectively.p = 0.128); however, the participants rated IADL capacity significantly higher than IADL performance (p < 0.01) (BADL performance and BADL capacity were not significantly different ( < 0.01) .p = 0.109); however, IADL capacity was rated significantly higher than performance (p < 0.01). Similarly, BADL performance and BADL capacity were not significantly different in participants with a CDR score of 1.0 (p = 0.715); however, IADL capacity was rated significantly higher than IADL performance (p < 0.01) , IADL performance and IADL capacity , and total ADL performance and total ADL capacity .ADL measures were also compared separately based on the participants’ CDR status. In participants with a CDR score of 0.5, there was no significant difference between BADL performance and BADL capacity ( < 0.01) . ParticiIn this study, we investigated ADL performance and capacity in patients with mild dementia. There was no gap between BADL performance and BADL capacity; however, the scores indicated that participants performed IADL less independently than their latent potential. Dementia is a progressive chronic disease that is intractable to significant improvements in mental and physical functioning. Environmental factors are one of the contextual factors in the International Classification of Functioning, Disability, and Health (ICF) . These fThere was no significant difference between the participants’ actual and latent abilities to perform BADL. In the mild dementia stage, BADL are less likely to decline in the first stage. Therefore, a gap between capacity and performance is considered unlikely to occur in BADL. On the other hand, IADL, which begin to decline from the mild dementia stage, are thought to have a gap due to environmental limitations ,11. One One notable point of this study is our successful identification of the gap between IADL performance and IADL capacity in persons with mild dementia by distinguishing between these concepts during the assessment. In addition, this finding supports Branch et al.’s assertions  that botThis study has certain limitations and unresolved issues that require further research. First, the study area was limited. The study covered day care centers in four regions of Japan. As a future developmental task, we would like to increase the number of participating regions to cover a wide range of cities with small to large populations to avoid geographical bias, and conduct the survey with greater statistical precision. Moreover, since the study focused on mild dementia, it cannot be generalized to people with moderate-to-severe dementia. Second, the findings cannot be generalized to all persons in residential areas because we excluded individuals living alone as control variables. Third, only a few participants qualified for the study. In the future, based on the findings of this study, it may be necessary to ensure a suitable number of participants, take into account the study area and living environment, and generalize the results to persons with dementia and their caregivers.Since people with mild dementia are in a situation where they are not able to fully demonstrate IADL capacity, there is ample room for intervention to improve their current situation. It is important for caregivers to make appropriate environmental adjustments in community life as these environmental interventions can contribute to improving the QOL of people with dementia.Since no curative treatment for dementia has yet been established, it is important to improve QOL and prevent progression from an early-stage dementia, and we hope that this study will contribute to effective non-pharmacological interventions.


GJB2 gene (encoding Connexin26(Cx26)) are the most common cause of hereditary deafness, accounting for about a quarter of all cases. Sensory epithelial damage is considered to be one of the main causes of deafness caused by GJB2 gene mutation. Dexamethasone (DEX) is widely used in the treatment of a variety of inner ear diseases including sudden sensorineural hearing loss (SSNHL), noise-induced hearing loss (NIHL), and deafness caused by ototoxic drugs. Whether DEX has a direct therapeutic effect on hereditary deafness, especially GJB2-related deafness, remains unclear. In this study, we revealed that DEX can effectively prevent hair cell death caused by oxidative stress in cochlear explants. Additionally, two distinct Cx26-null mouse models were established to investigate whether systemic administration of DEX alleviate the cochlear sensory epithelial injury or deafness in these models. In a specific longitudinally Cx26-null model that does not cause deafness, systemic administration of DEX prevents the degeneration of outer hair cells (OHCs) induced by Cx26 knockout. Similarly, in a targeted-Deiter’s cells (DCs) Cx26-null mouse model that causes deafness, treatment with DEX can almost completely prevent OHCs loss and alleviates auditory threshold shifts at some frequencies. Additionally, we observed that DEX inhibited the recruitment of CD45-positive cells in the targeted-DCs Cx26-null mice. Taken together, our results suggest that the protective effect of dexamethasone on cochlear sensory epithelial damage and partially rescue auditory function may be related to the regulation of inner ear immune response in Cx26 deficiency mouse models.Mutations in the  Hereditary hearing loss is the most common hereditary disease that affects every 1.4/1000 newborns worldwide –4. HowevGJB2 gene is the most common cause of hereditary deafness, accounting for about a quarter of hereditary deafness in different populations [Mutation in the ulations , 7. Connulations , 9. Gap ulations –12. Cochulations , 14. In ulations . Howeverulations . In addiulations . In thisulations . TherefoGlucocorticoids (GCs) are widely used for a variety of inner ear diseases, such as SSNHL, NIHL, ototoxic drug-induced hearing loss, and Meniere’s disease –23. DexaIn this study, a cochlear explant model of oxidative damage induced by glucose oxidase (GO) was established to investigate the therapeutic effect of DEX on oxidative damage to the cochlear sensory epithelium. Based on the evidence that dexamethasone has anti-oxidant, anti-inflammatory, and anti-apoptotic effects, we hypothesized that DEX may protect against the cochlear sensory epithelium injury or hearing loss induced by Cx26 deficiency. Two distinct Cx26-null mouse models were established to test our hypothesis. One line was DC-targeted Cx26-null mice, which exhibit rapid hair cell loss and deafness . The othf/f mice were crossed with Fgfr3iCreERT2 or Lgr5CreER mice to obtain Cx26f/f; Fgfr3iCreERT2 and Cx26f/f; Lgr5CreER mice. All mice were injected with tamoxifen  subcutaneously at P0 and P1  [f/f; Lgr5CreER mice, Cx26 of the third row of DCs was successfully knocked out [f/f; Fgfr3iCreERT2 line was used to establish a targeted DCs and PCs Cx26-null mouse model [All experiments were approved by the Committee of Tongji Medical College, Huazhong University of Science and Technology. Cx26ve days) , 36. In se model .All mice were raised in the specific-pathogen-free Experimental Animal Center of Huazhong University of Science and Technology. The animals were housed at 22 ± 1 °C under a standard 12 h light/dark cycle and were allowed free access to water and a regular mouse diet.2 overnight before each treatment. On the following day, the control group aswas cultured with the fresh medium without any drugs, GO group was exposed to 160 U/L glucose oxidase, GO + dexamethasone group was exposed to the same concentration of GO together with 100 μg/mL dexamethasone, or dexamethasone group was exposed to 100 μg/mL dexamethasone. All cochlear explants were incubated for 24 h and harvested for hair cell counting. Cochlear explants after GO treatment were washed in PBS and incubat with 10 mM DCFH-DA  in serum-free DMEM for 30 min.The preparation of the tail collagen gel matrix has been described in detail in our previous studies . BrieflyClinically, systemic dexamethasone has demonstrated conclusive benefits in reversing SSNHL despite considerable number of potential side effects. To test the protective effect of dexamethasone on the targeted-cell Cx26-null models. Mice in the dexamethasone intervention group were subcutaneously injected with dexamethasone every two days for eight consecutive days. Two concentrations of DEX (3 mg/kg and 5 mg/kg) were tested in our preliminary study, based on the literature , 39. Then = 5) were measured by ABR at P18. The detailed methods for the ABR test have been described in detail in our previous publications [The hearing thresholds of each group , the flattened cochlear preparations or cochlear explants were incubated in a blocking solution of 5% Bovine Serum Albumin, followed by incubation with primary antibodies: polyclonal rabbit anti-myosin7a antibodies , polyclonal goat anti-sox2 antibodies , polyclonal rabbit anti-Cx26 antibodies  and goat anti-CD45 polyclonal antibody  diluted in PBS overnight at 4 °C. The samples were washed three times with PBST and then incubated with secondary fluorescent antibodies  for 2 h at room temperature. Nuclei and F-actin staining were labeled with DAPI and phalloidin  for 10 min. The samples were visualized under a laser scanning confocal microscope .P < 0.05 was considered statistically significant.In vitro and in vivo experiments, each group contained at least four independent samples. All data are shown as the mean ± SD, Statistical analyses were conducted using GraphPad Prism  and SPSS software . One-way ANOVA followed by a Dunnett multiple comparisons test was used when there was only one factor. Two-way-ANOVA multiple comparisons test was used when two factors were involved. n = 5) showed that the survival rate of IHCs were 62.96 ± 5.48, 59.45 ± 9.13, and 57.98 ± 12.0% in the apical, middle, and basal turns, respectively , NIHL, ototoxic drug-induced hearing loss, as well as hereditary hearing loss. Recently, DEX was reported to act as a slow-acting free radical cleaner and has been shown to ameliorate hair cell damage induced by various exogenous stresses . In thisely Fig. . In contn = 4) showed that approximately 27.5 ± 2.60% of OHC3 was degenerated in the specific longitudinally Cx26-null group at P15 . Our results indicate that the earliest degeneration of OHCs occurred at P14–P15, suggesting that the window of time for hair cell injury intervention should be earlier than this time point.Our previous study showed that degeneration of OHCs was observed in multiple Cx26-null models at P18. To explore the appropriate treatment window, we dynamically observed hair cell survival in different Cx26-null mouse models Fig. . There wP15 Fig. . AdditioP15 Fig. . The whif/f; Lgr5-CreER mice were injected with tamoxifen to establish a specific longitudinally Cx26-null mouse model  had lost their Cx26 expression  showed that approximately 49.0 ± 4.58% OHC3 loss was observed in the specific longitudinally Cx26-null group at P18, while the loss of hair cells was significantly reduced in DEX and NAC treatment groups . There is almost no statistical difference in hearing between the Cx26-null+DEX group and the control group at P18. In addition, hair cells were labeled with Myosin7a by immunofluorescence staining. In the control group and the DEX alone treatment group, there was no obvious OHC loss at P18  showed that the survival rate of OHCs in the basal turn increased from 48.34 ± 4.52 to 88.22 ± 1.91% in the DEX treatment group . These results demonstrate that treatment with DEX prevents OHC loss and alleviates auditory threshold shifts at certain frequencies in the DC-targeted Cx26-null mouse model.Next, a DC-targeted Cx26-null mouse model was established to explore the protective effects of DEX Figs. , 5A. In P18 Fig. . ConsistP18 Fig. . In lineP18 Fig. . QuantifP = 0.0032, Fig. 2 vs 382.52 ± 55.29 mm2, P = 0.00468, Fig. Since our previous study has reported that the macrophage-related immune response is involved in the pathological process of cochlear sensory epithelium damage induced by Cx26 knockout . The numGjb2 mutation-related hereditary deafness, in which sensory epithelial damage is considered to be a recognized cause of deafness [Gjb2-related cochlear cell death. Besides, our preliminary study found that mouse pups older than 10 days could tolerate 5 doses of DEX injections, and this total dose of DEX causes death in pups less than 1 week old. It has been reported that deletion of Cx26 contributes to a diminished antioxidant defense system in the cochlea of Gjb2 knockout mice, indicating that accumulation of ROS might be involved in sensory epithelial damage [In the clinic, GJB2 mutations cause various auditory phenotypes, including profound sensorineural hearing loss or late-onset progressive hearing loss. This diversity of clinical phenotypes implies that hearing loss induced by Cx26 mutations has various underlying deafness mechanisms. Recent studies have shown that multiple pathological mechanisms are involved in deafness . In the deafness , 49. An l damage . Studiesl damage . In vitrl damage . In thisGjb2-related deafness, we established a targeted DC Cx26-null model with both OHC death and deafness. In this line, the location of OHC death was matched with the frequency of hearing loss, suggesting that the hearing loss in this model was mainly caused by OHC death. Our results showed that systemic DEX administration could almost completely prevent OHCs loss and deafness in the short term. So far, how Cx26 knockout of SCs leads to HC death remains unclear. Although we do not know the specific mechanism by which DEX antagonizes OHC death, it indicated that it is a feasible method to save Gjb2-related deafness based on protecting OHC death.To further verify the effectiveness of DEX in treating Gjb2-related hereditary deafness is significantly different from that of other types of deafness [Gjb2-related cochlear sensory epithelial injury and deafness. In this study, we found that treated with DEX significantly inhibited the recruitment of CD45-positive cells in the targeted-DCs Cx26-null mice, suggesting that the protective effect of DEX on OHC injury may be related to the inhibition of inner ear immune response. Our findings provide evidence for the immune response as promising new therapeutic targets for the prevention of Gjb2-related hearing loss. More drugs based on inhibition of inflammation in the inner ear could be tried to treat Gjb2-related deafness and cochlear sensory epithelial damage.There is growing evidence that immune responses are involved in a variety of acute or chronic inner ear diseases including hearing loss caused by acoustic injury, ototoxicity, toxins, and cochlear implantation. In recent years, treatments targeting inflammation have been shown to improve a variety of inner ear diseases. During these cochlear epithelium damage processes, infiltrating or mature tissue macrophages are the main immune response cells, several studies have demonstrated distinct roles for macrophages including antigen presentation, phagocytosis of cellular fragments, and production of inflammatory molecules , 51. Studeafness , 57. We Gjb2-related hearing loss. Yu et al. inoculated viral vectors into the scala media of conditional Cx26 knockout mice to reconstruct cochlear Cx26 expression, and extensive gap junctions among the supporting cells were successfully reconstructed. Degeneration of HCs and SGNs in conditional Cx26 knockout mice was significantly reduced following exogenous connexin26 expression. Unfortunately, there was no significant improvement in hearing [GJB2 mutation needs to be further studied.Up to now, there is still no effective treatment based on the mechanism of  hearing . Our stuGjb2-related sensory epithelial death, and antioxidants can partially block HC death. In the targeted-DC Cx26-null model, DEX is very effective in blocking HC death and restoring hearing in the short term. DEX may save Gjb2-associated HC death and hearing through a variety of pathways. These new findings have profound implications for the intervention in hereditary deafness caused by Cx26 mutations and show that a more comprehensive understanding of this common hereditary deafness is important and urgent.In conclusion, Cx26-null mice had an optimal deafness treatment window of about one week before the onset of sensory epithelial injury. Oxidative stress injury is an important cause of checklist


School-aged children may experience hearing loss and emotional problems. Previous studies have shown a bidirectional relationship between hearing loss and emotional problems in the elderly population, and we aimed to analyze the association between hearing thresholds and emotional problems in school-aged children.Based on the Beijing Child Growth and Health Cohort (PROC) study, the hearing screenings were conducted in November 2019 using pure tone audiometry. A total of 1,877 parents completed the Strengths and Difficulties Questionnaire (SDQ) to assess children's emotional and behavioral status. We used generalized linear regression analysis to assess the potential association of emotional problems with hearing thresholds, based on multiple imputed datasets with a sample size of 1,914.CI: 0.08–0.40), and right ear average hearing thresholds . Conduct problems, hyperactivity/inattention, peer problems, and prosocial behaviors had no association with the pass rate of the hearing screening. Regarding emotional symptoms, boys with many fears and who are easily scared coincided with increased right ear average hearing thresholds . Girls having many worries, frequently feeling unhappy and downhearted were positively associated with left and right ear average hearing thresholds, respectively .The overall pass rate of hearing screening was 91.5%. The abnormal rate of SDQ total difficulties was 55.8%. Emotional symptoms were positively associated with left ear average hearing thresholds (β = 0.24, 95%The co-occurrence of hearing problems and emotional problems of children aged 6–8 in Beijing attracts attention. It is important to address undiscovered hearing loss and emotional problems from the perspective of comorbidity driving factors. Hearing loss in children is becoming an urgent public health issue. For school-aged children, newly onset hearing loss remains a matter of great concern as one of the most common developmental disorders, and its prevalence increases till adulthood due to the additions of late-onset, late identified, and acquired hearing loss .The WHO estimates that approximately 34 million children worldwide live with hearing loss in 2021  assessment , 21. ChiCortical hemispheric asymmetries may be present in early childhood, it means the hemispheric superiority dominates in language processing . With fuThis study aimed to understand the hearing thresholds of children aged 6–8 years during their transition from kindergarten to primary school, and analyze the impact of emotional problems on hearing thresholds.The participants aged 6–8 years were recruited from the Beijing Child Growth and Health Cohort (PROC) Study, launched in October 2018 in six non-boarding primary schools in Beijing . All parThe hearing screenings were conducted in soundproof classrooms of each school from November 4-8, 2019. Before the screening, children were given a nasopharyngoscopy and otoscopy by an otolaryngologist to exclude possible hearing loss due to cerumen impaction and nasopharyngeal problems such as nasal obstruction. Children with cerumen impaction were included in this study after cerumen impaction was removed by the otolaryngologist. We exclude children who wore hearing aids or had otitis media because they were not suitable for hearing screening, and 1,839 of 1,914 children were screened (96.1%) except those who were absent on screening day.Pure tone audiometry was conducted by the Madsen Xeta screening audiometer  and the ambient noise was <45 dB A. The audiologists conducted the tests and explained the notes about the screening before the start of the test, to ensure that children understood the test procedure. The child was tested with right ear first   and raisAccording to the American Academy of Audiology Childhood Hearing Screening Guidelines, a pure tone sweep was conducted at 1,000, 2,000, and 4,000 Hz and 20 dB HL was used as MHL, a non-response to any frequency in either ear was considered a failure of the screening. The test sound can be given multiple times at each frequency in each ear, up to four times. For all children participating in the screening, a threshold test is performed using the “10 down, 5 up” method to determine a specific threshold as continuous measures to quantify the degree of hearing loss and those who failed were recommended for a referral to a clinical service for further diagnosis or treatment. The procedure and results of further diagnostic test were shown in the We conducted a baseline survey among the 1,914 children's parents from November 2018 to May 2019, and 1,910 parents responded (99.8%). Taking out 33 responses of incomplete answers, a total of 1,877 completed questionnaires were analyzed (98.3%).We used the parental version of the Strengths and Difficulties Questionnaire (SDQ) to assess the emotional and behavioral status of children, including 5 scales: (a) emotional symptoms; (b) conduct problems; (c) hyperactivity/inattention; (d) peer problems; and (e) prosocial behaviors. Each scale includes 5 questions and each question has three options, “not true”, “somewhat true” and “certainly true”. The options were given different scores “0”, “1” and “2” according to the meaning of each question, and the scores were calculated in subtotal and total. We categorized scores for each scale into 3 groups of “normal”, “borderline” and “abnormal”. Each group had different cutoff values, and the instruction of the questionnaire was followed per the SDQ website .p < 0.05 was considered statistically significant.We used Statistical Analysis System V.9.4  software for statistical analysis. Multiple imputation was used to process the missing data, and 7 imputed datasets were obtained. We used means ± standard deviation to describe continuous variables for normal or proximate to a normal distribution, and used number (%) to describe categorical variables. Generalized linear regression was used to analyze the potential association of the emotional problems on the mean thresholds of the right and left ears, unadjusted and adjusted for child sex, age, BMI, and parental education , 32. Furp < 0.001). The girls' maternal educational attainment is higher than the boys' (p = 0.018). The abnormal rate of SDQ in total difficulties reported by parents was 55.8%, with no differences between the sexes . The abnormality rates were, from highest to lowest, 17.2% for peer problems, 9.3% for emotional symptoms, 9.0% for conduct problems, 8.3% for hyperactivity/inattention, and 0.0% for prosocial behaviors. The borderline rates were sequential, 16.8% for peer problems, 14.2% for prosocial behaviors, 14.0% for conduct problems, 10.2% for hyperactivity/inattention, and 10.1% for emotional symptoms. The normal rates rank as, 85.8% for prosocial behaviors, 81.5% for hyperactivity/inattention, 80.6% for emotional symptoms, 77.1% for conduct problems, and 66.0% for peer problems. Except for the conduct problems (p = 0.12), the reporting rates for the remaining four problems differed between sexes (p < 0.05). Girls presented more emotional symptoms and boys presented more hyperactivity/inattention, peer problems and prosocial behaviors  was 6.6  years. Among them, 956 were boys. The boys' BMI is higher than the girls' (ehaviors .p < 0.05). And the hearing thresholds of the left ear at 4,000 Hz were marginally significant (p = 0.052) between sexes , and the right ear average hearing thresholds . Adjusting sex, age, BMI of children, and parental education, the association with the left ear average hearing thresholds was similar, and the association with right ear average hearing thresholds was marginally significant . Conduct problems, hyperactivity/inattention, peer problems, and prosocial behaviors did not have a significant impact on the hearing threshold . For girls, having many worries was positively associated with the left ear average hearing thresholds  and was marginal significant on increased right ear average hearing thresholds . Often being unhappy and downhearted was positively associated with the right ear average hearing thresholds   .We found that the hearing screening failure rate of children was 8.5% among children aged 6–8 years in Beijing, China, and the overall abnormal rate of SDQ in total difficulties score categorization reported by parents was 55.8%. The emotional symptoms were positively associated with the left and right ear average hearing threshold and presented a sex discrepancy.The hearing screening failure rates of children varied significantly across the globe. A Poland study which also used 20 dB HL as MHL showed that the estimated prevalence of hearing loss among 5,029 children aged 6–7 years was 11.6% . And othWe found that hearing thresholds differed by sex except for the hearing thresholds of the left ear at 1,000 Hz, with higher thresholds for girls than for boys, but no sex difference in screening failure rates, which is consistent with the findings of a study on preschool children . This reGlobally, an increasing number of children and adolescents suffer from hearing loss. Due to different characteristics of hearing loss , the prevalence of hearing loss increases throughout school age . A meta-This study found that the abnormal rate of SDQ total difficulties reported by parents was 55.8%, 57.4% for boys and 54.1% for girls. It is worth noting that compared to other studies, the children in this study had more severe emotional and behavioral problems. A study using the SDQ to screen for emotional and behavioral problems assessed 596 families among children aged 7–8 years and showed that the prevalence of emotional and behavioral problems was 30.0% for boys and 28.2% for girls. Hyperactivity/inattention was more prevalent among boys, which was consistent with this study . For chiChildren are only able to experience and perceive emotions when their brains develop a specific capacity . This isOne who pass the hearing screening does not mean that he or she has normal hearing certainly. People with higher hearing thresholds can manifest hearing impairment, communication difficulties, etc., even within the normal range , 65. It Our study has apparent strength. It is the first study to focus on the potential association of school-aged children's emotional problems with hearing using a prospective cohort study. There are also several limitations. The hearing screening and SDQ questionnaire surveys are cross-sectional at the baseline, so the cause-and-effect temporal relationships cannot be inferred definitively and in the future, we could continue to verify such association longitudinally. As the screening was performed in children aged 6–8 years in Beijing, the findings cannot be generalized to the entire childhood nor in other geographical areas. But it expanded knowledge in understanding children aged 6–8 years. The questionnaires were completed by parents based on their observations of the children for screening purposes of the prevalence of emotional and behavioral problems, not for diagnosis. Generally, SDQ is reliable and we can use it to identify children with a tendency for emotional and behavioral problems.In conclusion, hearing loss and emotional problems of school-aged children in Beijing are cause for concern. We emphasized the necessity and importance of regular audiological monitoring in school settings and call for the attention to undiscovered hearing loss as well as emotional problems. This study is innovative in its exploration of the interplay of emotional problems with hearing status in children. It is expected that we can inform school health policy to address both challenges in the near future.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Ethics Committee of Capital Medical University (No. 2018SY82). Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.YH is the principal investigator of the present study and conceptualized and designed the study. WW performed nasopharyngeal and otoscopy examination. LH, HE, XZ, CW, YY, HX, NA, ML, and JZ carried out the hearing screening. HX and NA conducted data analysis. HX and YH drafted the manuscript. YH, WW, LH, HE, XZ, CW, YY, HX, NA, ML, and JZ interpreted findings, edited, and revised the manuscript. All authors had final approval of the submitted and published versions and agreed to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.The study was supported in part by the National Natural Science Foundation of China , the Beijing Natural Science Foundation (7202009) and the Capital Health Research and Development of Special (2022-2-1092). The sponsors have no role in study design, collection, analysis, and interpretation of data, the writing of the report, and in the decision to submit the paper for publication.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


We present a case of a 42-year-old female living with poorly controlled diabetes who presented with a nine-month evolution of ataxic gait, reduced motor and sensitive function of lower and upper limbs, and postural anesthesia of fingers, feet, and toes. Deep tendon reflexes were abolished in the lower limbs and markedly diminished in the upper limbs. Cerebrospinal fluid (CSF) analysis showed a high protein level, and both imaging and serologic studies were normal. Although she had a previous electrophysiologic study showing distal symmetric polyneuropathy (DSPN) with an axonal lesion, nerve conduction studies were repeated, and a diagnosis of chronic inflammatory demyelinating polyneuroradiculopathy (CIDP) was made. According to the state of the art, intravenous immunoglobulin (IVIg) was started. The patient’s Inflammatory Neuropathy Cause and Treatment (INCAT) score and Medical Research Council (MRC) Sum Score both improved after two cycles. Unfortunately, symptoms quickly recurred, and corticosteroids were introduced to try to delay symptom recurrence, although it worsened diabetes control. Later, IVIg was stopped due to nephrotic syndrome, and immunosuppression was initiated. CIDP is a potentially treatable disease, but the diagnosis must be made as soon as possible to start therapy and reduce sequelae. Neuropathy in patients living with diabetes is common, but patients must be monitored closely to enable a correct diagnosis and adequate treatment. Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is an acquired immune-mediated disorder of peripheral nerves . AlthougA 42-year-old female presented to the outpatient internal medicine clinic with a nine-month history of progressive hypoesthesia and muscle weakness of the upper and lower limbs progressing from distal to proximal regions and ataxic gait. She had been living with poorly controlled diabetes with microvascular complications  and controlled hypertension. She was medicated with detemir insulin, metformin, vildagliptin, and enalapril. Family history was not relevant, with no history of neurologic disease.Since the onset of symptoms, she had been observed at the emergency department for a vertigo episode where a cerebral computed tomography (CT) revealed no signs of ischemic nor hemorrhagic vascular lesion and no cerebral mass. Benign paroxysmal positional vertigo (BPPV) was excluded, and cervical CT was normal. All examination results are presented in Table Neurologic examination revealed no alteration of mental status or cranial nerves. She presented with an ataxic gait, being unable to walk without support or walk on her heels or toes. The Romberg test was positive, and minor terminal dysmetria was noted on the finger-nose test with a normal heel-knee test. Lower limb deep tendon reflexes were abolished and markedly diminished in the upper limbs. Motor function was altered with a positive Mingazzini test and diminished muscle strength with distal predominance being observed. Interosseous muscle atrophy was noted in both hands and feet. Sensory function examination of the upper limbs documented tactile hypoesthesia and hypoalgesia with postural hypoesthesia of the fingers. On the lower limbs, tactile anesthesia, analgesia, and vibratory hypoesthesia with postural anesthesia of feet and toes were present. Lhermitte’s sign was also present. At that point, posterior cord syndrome and peripheric neuropathy were considered.Borrelia infection, and autoimmunity for neurologic and systemic diseases were negative. Protein electrophoresis and immunofixation were normal, excluding polyneuropathy, organomegaly, endocrinopathy, monoclonal protein, and skin change (POEMS) syndrome. Although there were previous nerve conduction studies (NCS) that showed axonal sensitive polyneuropathy, NCS was repeated due to new and rapidly progressive neurologic signs and symptoms. NCS revealed features of CIDP in more than two different nerves in both lower and upper limbs. At this point, a definitive typical CIDP diagnosis was confirmed, meeting all clinical criteria, electrophysiologic criteria in more than two nerves, and with cerebrospinal fluid protein elevation as supportive criteria.Medullar and cerebral magnetic resonance (MRI) showed no traumatic abnormalities nor signs of vasculitis, and there was no history of radiation. Cerebrospinal fluid (CSF) showed hyperproteinorachia with albuminocytological dissociation. The Tibbling index was normal, and there were no oligoclonal bands. Somatosensory evoked potentials showed no alterations. Laboratory blood analysis confirmed a poorly controlled diabetes with glycated hemoglobin of 12.5%. Thyroid hormones were normal, and there was no vitamin B12 deficiency. Serologies for HIV, syphilis, According to the European Academy of Neurology/Peripheral Nerve Society (EFNS/PNS) guidelines, intravenous human immunoglobulin (IVIg) was started at a dose of 2 g/kg for five days with significant clinical improvement. Intensive glycemic control and neuromodulator pain therapy had already been implemented.The patient was discharged after completing the first IVIg and was re-evaluated two weeks later. Due to significant clinical improvement after the first cycle of IVIg and after consulting with the neurology department, the same dose was repeated one month later. After the first two IVIg cycles, the Inflammatory Neuropathy Cause and Treatment (INCAT) disability scale score went from 6 to 3, and the Medical Research Council (MRC) Sum Score went from 42 to 54, a satisfactory response to the IVIg and another supportive criterion of CIDP diagnosis. Treatment was continued, according to EFNS/PNS guidelines, with a scheme of IVIg 1 g/kg for two days every three weeks.Symptoms recurred soon after each infusion despite good glycemic control. After the fourth cycle of IVIg, glucocorticoids were introduced to control neurologic symptoms between IVIg infusions. Unfortunately, the patient developed nephrotic syndrome, and IVIg had to be suspended. Immunosuppression was started with azathioprine with very little result. Treatment of neuropathic pain was optimized, improving the patient’s quality of life.Diagnostic criteria were reviewed in 2021, and CIDP is now classified as typical CIDP and CIDP variants . TypicalDiagnosis may be challenging, and CIDP is often underdiagnosed or misdiagnosed as presentation varies widely and diagnostic criteria are only clinical and electrophysiologic ,6. OtherA retrospective study with 60 patients with definitive diagnoses of CIDP in a reference center showed that 68.3% had an alternative diagnosis, the most common being GBS (23.3%), often misdiagnosed in acute CIDP onset. Non-GBS alternative diagnoses 45%) were mainly misdiagnosed as genetic neuropathy, diabetic neuropathy, and chronic idiopathic axonal polyneuropathy [% were maPatients with CIDP often have comorbidities that may influence diagnosis and treatment choice, such as diabetes mellitus . AlthougDistal symmetric polyneuropathy (DSPN) is the most common diabetic neuropathy . It presThe first-line recommended treatment for CIDP is immunoglobulin (intravenous (IVIg) for induction and subcutaneous for maintenance), corticosteroids, or plasma exchange. EFNS/PNS guidelines advocate the first two as equally effective and recommend IVIg over plasma exchange due to ease of administration. Although there are no specific recommendations for CIDP treatment in people with diabetes, poor glycemic control in these patients is very common and usually results in IVIg use over corticosteroids .Response rates to CIDP treatment seem to be similar in patients with and without diabetes, although underlying DSPN will limit the response, and full normalization of function and strength might not occur in patients with diabetes .Although the CIDP-DM association has yet to be clarified, people who live with diabetes, particularly patients with diabetic neuropathy, should be monitored closely. In these patients, previous neurologic symptoms may be a confounding bias, resulting in delayed CIDP diagnosis and consequently worse outcomes.CIDP treatment should be implemented as soon as possible, alongside glycemic control and nerve pain control, to reduce symptoms and improve patients’ quality of life.


The study aimed to examine the effects of hearing aids on cognitive function in middle-aged and older adults with hearing loss.PubMed, Cochrane Library, and Embase were searched for studies published before 30 March 2022. Randomized controlled trials (RCTs) and non-randomized studies of interventions (NRSIs) were included in the search. Restriction was set on neither types, severity, or the time of onset of hearing impairment nor cognitive or psychiatric statuses.Two independent reviewers extracted data and assessed the study quality of RCTs. Cognitive function outcomes were descriptively summarized and converted to standardized mean difference (SMD) in the meta-analysis. Meta-analysis was conducted in RCTs. Sub-group analyses were conducted by cognitive statuses, psychiatric disorders, and cognitive domains.n = 339) and 10 NRSIs (n = 507). Groups were classified as subjects without dementia or with normal global cognition, subjects with AD or dementia, and subjects with depressive symptoms. For subjects without dementia, improvements were found in global cognition, executive function, and episodic memory. For subjects with depressive symptoms, improvements were found in immediate memory, global cognition, and executive function. No improvement was found in subjects with AD or dementia. In total, four RCTs were included in the meta-analysis. For subjects without dementia  and those with AD, no significant effect was found . For subjects without dementia, no significant effect was found in language  or general executive function . Further sub-group analysis found no significant effect in executive function  or processing speed .A total of 15 studies met the inclusion criteria, including five RCTs (Hearing aids might improve cognitive performance in domains such as executive function in subjects without dementia. The effects on subjects with depressive symptoms remained unclear. No improvement was found in subjects with AD or dementia. Long-term RCTs and well-matched comparison-group studies with large sample sizes are warranted.https://www.crd.york.ac.uk/PROSPERO/, identifier: CRD42022349057. Nowadays, the rate of hearing loss presents an increasing trend. For example, in China, according to the research by Hu et al. , the preAlthough the mechanism underlying hearing loss and cognitive decline is not yet clear, indeed, studies suggesting improvements in cognition with hearing loss management have been conducted, possibly through enhancing communication and improving the quality of life  guidelines : (1) PubMed, (2) Cochrane Library, and (3) Embase. Keywords included hearing aids, Alzheimer's disease, dementia, mild cognitive impairment, and cognitive function. Details of the search strategy and results are given in The inclusion criteria for the articles are as follows: (1) prospective, original articles of randomized controlled trials (RCTs) and non-randomized studies of interventions (NRSIs), which included non-randomized controlled trials (non-RCTs) and pretest–posttest studies using a within-subject design (without control or comparison groups); (2) full-text articles published in English only; (3) articles including subjects aged ≥ 45 years with hearing impairment; (4) those with no restriction on the assessment of hearing status; (5) those with full inclusion of hearing loss samples ; (6) articles with no restriction on cognitive status or other psychiatric disorder of subjects; and (7) those including hearing aids as an intervention in experimental groups. We excluded non-longitudinal studies and observational studies.Two independent authors (Yang Z and Teng Y) screened for the eligible studies and conducted data extraction. The corresponding author (Shi J) acted as an arbitrator for the final decision if a consensus could not be reached. Data from studies that investigated hearing loss in adults with different cognitive statuses or psychiatric disorders were grouped. Cognitive domains were subdivided as described in the study by Lezak et al. : global The Cochrane protocol for assessing the risk of bias (RoB2) was used for the quality assessment of RCTs. RoB2 consists of five domains: bias arising from the randomization process, bias due to deviations from intended interventions, bias due to missing outcome data, bias in the measurement of the outcome, and bias in the selection of the reported result .2 and s22 were the variances of each study.The meta-analysis was restricted to RCTs  ] ] . Studiesp = 0.53; )  and found that hearing aids were not significantly more effective than the control condition where subjects received successful aging intervention at a 6-month follow-up    .p = 0.53; ]  and Boston Naming Test) and found that hearing aids were not significantly more effective than the control condition where subjects received successful aging intervention at a 6-month follow-up   ] .p = 0.71; )  part A and B), processing speed (measured by the Digit Symbol Substitution Test and Pattern Comparison Processing Speed Test), attention (measured by the Flanker Test), and working memory (measured by the List Sorting Working Memory Test) and found that hearing aids were not significantly more effective than the control condition where subjects received the successful aging intervention or no treatment at 6-month follow-up    .For further research, the sub-group analysis of single executive function was conducted; the results showed that hearing aids were not significantly more effective on executive function  and processing speed  . The resp = 0.43; )   (Nguyen 2 = 0%]) . The resThe risk of bias of the included RCTs is shown in Of three non-RCTs that were included, one was conducted in Germany . However, in the research by Doherty and Desjardins       , Qihuang Scholar Foundation (China), and the collaborative research project of Beijing University of Chinese Medicine (BZY-JMZY-2022-002).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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.