EBF1 Limits the Numbers of Cochlear Hair and Supporting Cells and Forms the Scala Tympani and Spiral Limbus during Inner Ear Development.

Early B-cell factor 1 (EBF1) is a basic helix-loop-helix transcription factor essential for the differentiation of various tissues. Our single-cell RNA sequencing data suggest that Ebf1 is expressed in the sensory epithelium of the mouse inner ear. Here, we found that the murine Ebf1 gene and its protein are expressed in the prosensory domain of the inner ear, medial region of the cochlear duct floor, otic mesenchyme, and cochleovestibular ganglion. Ebf1 deletion in mice results in incomplete formation of the spiral limbus and scala tympani, increased number of cells in the organ of Corti and Kölliker's organ, and aberrant course of the spiral ganglion axons. Ebf1 deletion in the mouse cochlear epithelia caused the proliferation of SOX2-positive cochlear cells at E13.5, indicating that EBF1 suppresses the proliferation of the prosensory domain and cells of Kölliker's organ to facilitate the development of appropriate numbers of hair and supporting cells. Furthermore, mice with deletion of cochlear epithelium-specific Ebf1 showed poor postnatal hearing function. Our results suggest that Ebf1 is essential for normal auditory function in mammals.

Mutations in PIH proteins MOT48, TWI1 and PF13 define common and unique steps for preassembly of each, different ciliary dynein.

Ciliary dyneins are preassembled in the cytoplasm before being transported into cilia, and a family of proteins containing the PIH1 domain, PIH proteins, are involved in the assembly process. However, the functional differences and relationships between members of this family of proteins remain largely unknown. Using Chlamydomonas reinhardtii as a model, we isolated and characterized two novel Chlamydomonas PIH preassembly mutants, mot48-2 and twi1-1. A new allele of mot48 (ida10), mot48-2, shows large defects in ciliary dynein assembly in the axoneme and altered motility. A second mutant, twi1-1, shows comparatively smaller defects in motility and dynein assembly. A double mutant mot48-2; twi1-1 displays greater reduction in motility and in dynein assembly compared to each single mutant. Similarly, a double mutant twi1-1; pf13 also shows a significantly greater defect in motility and dynein assembly than either parent mutant. Thus, MOT48 (IDA10), TWI1 and PF13 may define different steps, and have partially overlapping functions, in a pathway required for ciliary dynein preassembly. Together, our data suggest the three PIH proteins function in preassembly steps that are both common and unique for different ciliary dyneins.

In silico analysis of inner ear development using public whole embryonic body single-cell RNA-sequencing data.

The inner ear comprises four epithelial domains: the cochlea, vestibule, semicircular canals, and endolymphatic duct/sac. These structures are segregated at embryonic day 13.5 (E13.5). However, these four anatomical structures remain undefined at E10.5. Here, we aimed to identify lineage-specific genes in the early developing inner ear using published data obtained from single-cell RNA-sequencing (scRNA-seq) of embryonic mice. We downloaded 5000 single-cell transcriptome data, named 'auditory epithelial trajectory', from the Mouse Organogenesis Cell Atlas. The dataset was supposed to include otic epithelial cells at E9.5-13.5. We projected the 5000 â€‹cells onto a two-dimensional space encoding the transcriptional state and visualised the pattern of otic epithelial cell differentiation. We identified 15 clusters, which were annotated as one of the four components of the inner ear epithelium using known genes that characterise the four different tissues. Additionally, we classified 15 clusters into sub-regions of the four inner ear components. By comparing transcriptomes between these 15 clusters, we identified several candidates of lineage-specific genes. Characterising these new candidate genes will help future studies about inner ear development.

Light-Triggered, Non-Centrosymmetric Self-Assembly of Aqueous Arylazopyrazoles at the Air-Water Interface and Switching of Second-Harmonic Generation.

Trans-p-methoxy arylazopyrazole spontaneously forms non-centrosymmetric polar crystals, which reversibly undergo liquefaction upon photoisomerization to the cis-isomer. This liquid cis-isomer has a large electric dipole moment and is highly soluble in water (solubility up to ≈58 mM), which is remarkably higher than that of the trans-isomer (690 µM). Vis-light illumination of the aqueous cis-isomer generates macroscopically oriented, non-centrosymmetric crystals at the air-water interface. Polar crystals are also formed in sandwich glass cells (spacing, 20 µm) upon photo-induced crystallization of the liquid cis-isomer. The trans-crystals thus formed showed second harmonic generation (SHG) whose intensity is switched on/off in response to the photo-induced phase transition.

Enhanced Electric Polarization and Polar Switching of Dipolar Aromatic Liquids Confined in Supramolecular Gel Networks.

Dipolar aromatic liquids confined in the interstitial domains of chiral organogels show significantly enhanced electric polarization, as compared with those of pure liquids alone or organogels formed with nonpolar liquids. Intriguingly, nitrobenzene gels showed a supramolecular polar switching phenomenon; i.e., hysteresis in the polarization (P)-electric field (E) curves was observed for the gel above the melting point of the solvent. This indicates that the nitrobenzene molecules confined in the chiral nanofibrous gel networks exert macroscopic polarization whose direction is inversed depending on the direction of the external electric field. The anomalously enhanced electric polarization and polar switching phenomenon of supramolecular gels in varied solvents are scrutinized by the positive-up-negative-down (PUND) measurements, and the interactions between the gel nanofibers and the polar solvent molecules play crucial roles for the emergence of the polar switching phenomenon. This work presents for the first time that dipolar liquid molecules filling the interstitial space of supramolecular gels exhibit a significant confinement effect. It provides a new perspective to design electric-field-responsive soft materials based on the functional liquid domains confined in their porous networks.

Chlamydomonas DYX1C1/PF23 is essential for axonemal assembly and proper morphology of inner dynein arms.

Cytoplasmic assembly of ciliary dyneins, a process known as preassembly, requires numerous non-dynein proteins, but the identities and functions of these proteins are not fully elucidated. Here, we show that the classical Chlamydomonas motility mutant pf23 is defective in the Chlamydomonas homolog of DYX1C1. The pf23 mutant has a 494 bp deletion in the DYX1C1 gene and expresses a shorter DYX1C1 protein in the cytoplasm. Structural analyses, using cryo-ET, reveal that pf23 axonemes lack most of the inner dynein arms. Spectral counting confirms that DYX1C1 is essential for the assembly of the majority of ciliary inner dynein arms (IDA) as well as a fraction of the outer dynein arms (ODA). A C-terminal truncation of DYX1C1 shows a reduction in a subset of these ciliary IDAs. Sucrose gradients of cytoplasmic extracts show that preassembled ciliary dyneins are reduced compared to wild-type, which suggests an important role in dynein complex stability. The role of PF23/DYX1C1 remains unknown, but we suggest that DYX1C1 could provide a scaffold for macromolecular assembly.

Prognostic impact of incisional or excisional biopsy of cervical lymph node metastases of solid tumors.

OBJECTIVES: In performing an open biopsy of a neck mass, an incisional biopsy may increase the risk of cancer cell seeding and dissemination that, ultimately, worsens a patient's survival. The aim of this study was to compare the impact of incisional and excisional biopsies of cervical lymph node metastases of solid tumors on patients' survival. METHODS: A retrospective review was made of patients with cervical metastases of solid tumors who underwent an open biopsy for a diagnosis between 2005 and 2015. Sixty-four patients met the criteria out of 524 open biopsy cases undertaken during the period. Survival analyses were estimated from 33 cases whose initial symptoms were the presence of a neck mass, using two modes of biopsy: excisional and incisional. RESULTS: The 2-year overall survival rates in incisional and excisional biopsy groups were 65% and 43%, respectively, and 2-year disease-specific survival rates were 74% and 43%, respectively. The differences were not significant. For lung cancer or head and neck cancer subgroups, survival differences between incisional and excisional biopsy groups were also not significant. CONCLUSIONS: A carefully targeted physical examination and performing a fine needle aspiration are essential to establish a diagnosis for the etiology of an unknown neck mass. In performing an open biopsy, the effect of an incisional biopsy on patients' survival was no worse than that of an excisional biopsy, despite the latter being theoretically preferable.

A simple and fast approach for missing-wedge invariant classification of subtomograms extracted from filamentous structures.

Unsupervised classification of subtomograms extracted from cryo-electron tomograms is often challenging due to the presence of a missing wedge in tomographic data. Here, we propose a simple new approach to classify subtomograms extracted from cryo-electron tomograms of filamentous objects. This unsupervised classification approach uses the 1D projections of the subtomograms for classification and works independently of the orientations of the missing wedge. We applied this approach to subtomograms from eukaryotic cilia and successfully detected heterogeneity including structural polymorphism of dynein molecules.

Facial flushing after alcohol intake as a predictor for a high risk of synchronous or metachronous cancer of the upper gastrointestinal tract.

BACKGROUND: In Japan, there has been a lot of reports showing an association between facial flushing after light alcohol consumption and heterozygosity for inactive aldehyde dehydrogenase-2 (ALDH2). Persons with inactive ALDH2 may have a higher risk of alcohol-related oral, pharyngeal and esophageal cancers, compared with those with wild-type ALDH2. The purpose of this study was to examine whether flushers with oral or pharyngeal squamous cell carcinoma have an increased risk of synchronous or metachronous cancer of the upper gastrointestinal (UGI) tract. METHODS: A retrospective study was performed by medical chart review and through a questionnaire sent to 285 patients treated for oral and pharyngeal cancer. Responses were obtained from 150 patients (52.6%), who were classified as flushers or non-flushers, smokers (≥20 pack-year; 1 pack-years = number of cigarettes/20 per day) or non-smokers, and drinkers (≥14 units of alcohol consumption per week; 1 unit = 22 g) or non-drinkers. Relationships of these factors with occurrence of second primary cancers (SPCs) in the UGI tract were investigated. RESULTS: In Kaplan-Meier analysis, there was a significantly higher rate of SPC at 5 years in flushers and drinkers, but no relationship with smoking. In multivariate analyses, a history of flushing was significantly associated with SPC in the UGI tract (HR 2.64, 95% CI 1.25-5.52, P = 0.0109), but not with smoking or alcohol consumption. CONCLUSIONS: A simple interview on history of facial flushing after alcohol intake can be useful for identifying patients at high risk for synchronous or metachronous cancers of the UGI tract.

Functional and structural significance of the inner-arm-dynein subspecies d in ciliary motility.

Motile cilia play various important physiological roles in eukaryotic organisms including cell motility and fertility. Inside motile cilia, large motor-protein complexes called "ciliary dyneins" coordinate their activities and drive ciliary motility. The ciliary dyneins include the outer-arm dyneins, the double-headed inner-arm dynein (IDA f/I1), and several single-headed inner-arm dyneins (IDAs a, b, c, d, e, and g). Among these single-headed IDAs, one of the ciliary dyneins, IDA d, is of particular interest because of its unique properties and subunit composition. In addition, defects in this subspecies have recently been associated with several types of ciliopathies in humans, such as primary ciliary dyskinesia and multiple morphologic abnormalities of the flagellum. In this mini-review, we discuss the composition, structure, and motor properties of IDA d, which have been studied in the model organism Chlamydomonas reinhardtii, and further discuss the relationship between IDA d and human ciliopathies. In addition, we provide future perspectives and discuss remaining questions regarding this intriguing dynein subspecies.

Chlamydomonas dynein-preassembly-deficient mutants exhibit characteristic ciliary responses to viscous media.

Ciliary-dynein preassembly in the cytoplasm is critical for the assembly and movement of motile cilia, organelles that function under viscous conditions. Defects in preassembly often lead to a reduction in specific types of ciliary dyneins. Here, we investigated how environmental viscosity affects the motility of preassembly-deficient cilia in the alga Chlamydomonas. We found that, depending on the type of ciliary dynein deficiency, each Chlamydomonas mutant displays a characteristic phenotype in cell propulsion. Our results highlight not only the unique function(s) of each dynein species, but also the importance of functional coordination between dyneins for ciliary motility under viscous conditions.

Molecular differences between neonatal and adult stria vascularis from organotypic explants and transcriptomics.

Summary: The stria vascularis (SV), part of the blood-labyrinth barrier, is an essential component of the inner ear that regulates the ionic environment required for hearing. SV degeneration disrupts cochlear homeostasis, leading to irreversible hearing loss, yet a comprehensive understanding of the SV, and consequently therapeutic availability for SV degeneration, is lacking. We developed a whole-tissue explant model from neonatal and adult mice to create a robust platform for SV research. We validated our model by demonstrating that the proliferative behaviour of the SV in vitro mimics SV in vivo, providing a representative model and advancing high-throughput SV research. We also provided evidence for pharmacological intervention in our system by investigating the role of Wnt/ß-catenin signaling in SV proliferation. Finally, we performed single-cell RNA sequencing from in vivo neonatal and adult mouse SV and revealed key genes and pathways that may play a role in SV proliferation and maintenance. Together, our results contribute new insights into investigating biological solutions for SV-associated hearing loss. Significance: Hearing loss impairs our ability to communicate with people and interact with our environment. This can lead to social isolation, depression, cognitive deficits, and dementia. Inner ear degeneration is a primary cause of hearing loss, and our study provides an in depth look at one of the major sites of inner ear degeneration: the stria vascularis. The stria vascularis and associated blood-labyrinth barrier maintain the functional integrity of the auditory system, yet it is relatively understudied. By developing a new in vitro model for the young and adult stria vascularis and using single cell RNA sequencing, our study provides a novel approach to studying this tissue, contributing new insights and widespread implications for auditory neuroscience and regenerative medicine. Highlights: - We established an organotypic explant system of the neonatal and adult stria vascularis with an intact blood-labyrinth barrier. - Proliferation of the stria vascularis decreases with age in vitro , modelling its proliferative behaviour in vivo . - Pharmacological studies using our in vitro SV model open possibilities for testing injury paradigms and therapeutic interventions. - Inhibition of Wnt signalling decreases proliferation in neonatal stria vascularis.- We identified key genes and transcription factors unique to developing and mature SV cell types using single cell RNA sequencing.

Effects of θ High Definition-Transcranial Alternating Current Stimulation in the Anterior Cingulate Cortex on the Dominance of Attention Focus in Standing Postural Control.

Attention focus affects performance in postural control while standing, and it is divided into internal focus (IF) and external focus (EF). Each individual has a predominant attention focus, and research has revealed that the dominance of attention focus may be an acquired trait. However, the impact of non-invasive brain stimulation on attention-focus dominance remains unexplored in the current literature. Here, we examined the effect of high-definition transcranial alternating current stimulation (HD-tACS) on θ waves in the anterior cingulate cortex (ACC) on standing postural control tasks in an EF condition for IF- and EF-dominant groups. The effect of θ HD-tACS on the ACC differed between IF- and EF-dominant groups, and θ HD-tACS in the IF-dominant group decreased the performance of standing postural control under the EF condition. The forced activation of the ACC with θ HD-tACS may have conversely reduced the activity of brain regions normally activated by the IF-dominant group. Additionally, the activation of ACC prioritized visual information processing and suppressed the superficial sensory processing that is normally potentially prioritized by the IF-dominant group. These results highlight the importance of changing the type of rehabilitation and sports training tasks to account for the individual's dominance of attention focus.

Effects of neurofeedback training combined with transcranial direct current stimulation on motor imagery: A randomized controlled trial.

Introduction: Neurofeedback (NFB) training and transcranial direct current stimulation (tDCS) have been shown to individually improve motor imagery (MI) abilities. However, the effect of combining both of them with MI has not been verified. Therefore, the aim of this study was to examine the effect of applying tDCS directly before MI with NFB. Methods: Participants were divided into an NFB group (n = 10) that performed MI with NFB and an NFB + tDCS group (n = 10) that received tDCS for 10 min before MI with NFB. Both groups performed 60 MI trials with NFB. The MI task was performed 20 times without NFB before and after training, and µ-event-related desynchronization (ERD) and vividness MI were evaluated. Results: µ-ERD increased significantly in the NFB + tDCS group compared to the NFB group. MI vividness significantly increased before and after training. Discussion: Transcranial direct current stimulation and NFB modulate different processes with respect to MI ability improvement; hence, their combination might further improve MI performance. The results of this study indicate that the combination of NFB and tDCS for MI is more effective in improving MI abilities than applying them individually.

Stepwise fate conversion of supporting cells to sensory hair cells in the chick auditory epithelium.

In contrast to mammals, the avian cochlea, specifically the basilar papilla, can regenerate sensory hair cells, which involves fate conversion of supporting cells to hair cells. To determine the mechanisms for converting supporting cells to hair cells, we used single-cell RNA sequencing during hair cell regeneration in explant cultures of chick basilar papillae. We identified dynamic changes in the gene expression of supporting cells, and the pseudotime trajectory analysis demonstrated the stepwise fate conversion from supporting cells to hair cells. Initially, supporting cell identity was erased and transition to the precursor state occurred. A subsequent gain in hair cell identity progressed together with downregulation of precursor-state genes. Transforming growth factor ß receptor 1-mediated signaling was involved in induction of the initial step, and its inhibition resulted in suppression of hair cell regeneration. Our data provide new insights for understanding fate conversion from supporting cells to hair cells in avian basilar papillae.

Effects of neurofeedback on standing postural control task with combined imagined and executed movements.

Introduction: Motor imagery (MI) is a method of imagining movement without actual movement, and its use in combination with motor execution (ME) enhances the effects of motor learning. Neurofeedback (NFB) is another method that promotes the effects of MI. This study aimed to investigate the effects of NFB on combined MI and ME (MIME) training in a standing postural control task. Methods: Sixteen participants were randomly divided into MIME and MIME + NFB groups and performed 10 trials of a postural control task on an unstable board, with nine trials of MI in between. Electroencephalogram was assessed during MI, and the MIME + NFB group received neurofeedback on the degree of MI via auditory stimulation. A postural control task using an unstable board was performed before and after the MIME task, during which postural instability was evaluated. Results: Postural instability was reduced after the MIME task in both groups. In addition, the root mean square, which indicates the sway of the unstable board, was significantly reduced in the MIME + NFB group compared to that in the MIME group. Conclusion: Our results indicate that MIME training is effective for motor learning of standing postural control. Furthermore, when MI and ME are combined, the feedback on the degree of MI enhances the learning effect.

Reliability and validity of the Japanese movement imagery questionnaire-revised second version.

OBJECTIVE: Developing a Japanese version of the Movement Imagery Questionnaire-Revised Second Version (MIQ-RS) is essential for widespread evaluation and treatment based on motor imagery in physically disabled persons and patients in rehabilitation. This study aimed to investigate the reliability and validity of the Movement Imagery Questionnaire-Revised Second Version (MIQ-RS), which assesses motor imagery ability, by translating it into Japanese. RESULTS: This study enrolled twenty healthy participants (10 men and 10 women, mean age 21.17 ± 1.10 years). Reliability was examined for internal consistency using Cronbach's alpha coefficient. Spearman's rank correlation coefficient was used to examine the criterion-related validity of the MIQ-RS and the Kinesthetic and Visual Imagery Questionnaire (KVIQ-20). Results showed that Cronbach's alpha coefficients for the MIQ-RS were 0.81 and 0.82 for visual and kinesthetic imagery, respectively. Significant positive correlations were found between each visual and kinesthetic imagery score, and each total on the MIQ-RS and KVIQ-20 scores (r = 0.73, p < 0.01; r = 0.84, p < 0.01; r = 0.80, p < 0.01, respectively). This study suggests that the Japanese version of the MIQ-RS is a reliable and valid method of assessing motor imagery ability.

Repetitive Peripheral Magnetic Stimulation Combined with Motor Imagery Changes Resting-State EEG Activity: A Randomized Controlled Trial.

Repetitive peripheral magnetic stimulation is a novel non-invasive technique for applying repetitive magnetic stimulation to the peripheral nerves and muscles. Contrarily, a person imagines that he/she is exercising during motor imagery. Resting-state electroencephalography can evaluate the ability of motor imagery; however, the effects of motor imagery and repetitive peripheral magnetic stimulation on resting-state electroencephalography are unknown. We examined the effects of motor imagery and repetitive peripheral magnetic stimulation on the vividness of motor imagery and resting-state electroencephalography. The participants were divided into a motor imagery group and motor imagery and repetitive peripheral magnetic stimulation group. They performed 60 motor imagery tasks involving wrist dorsiflexion movement. In the motor imagery and repetitive peripheral magnetic stimulation group, we applied repetitive peripheral magnetic stimulation to the extensor carpi radialis longus muscle during motor imagery. We measured the vividness of motor imagery and resting-state electroencephalography before and after the task. Both groups displayed a significant increase in the vividness of motor imagery. The motor imagery and repetitive peripheral magnetic stimulation group exhibited increased ß activity in the anterior cingulate cortex by source localization for electroencephalography. Hence, combined motor imagery and repetitive peripheral magnetic stimulation changes the resting-state electroencephalography activity and may promote motor imagery.

Structural comparison of n-type and p-type LaAlO3/SrTiO3 interfaces.

Using a surface x-ray diffraction technique, we investigated the atomic structure of two types of interfaces between LaAlO3 and SrTiO3, that is, p-type (SrO/AlO2) and n-type (TiO2/LaO) interfaces. Our results demonstrate that the SrTiO3 in the sample with the n-type interface has a large polarized region, while that with the p-type interface has a limited polarized region. In addition, atomic intermixing was observed to extend deeper into the SrTiO3 substrate at the n-type interface compared to the p type. These differences result in distinct degrees of band bending, which likely contributes to the striking contrast in electrical conductivity between the two types of interfaces.

Regulation of ciliary motility: conserved protein kinases and phosphatases are targeted and anchored in the ciliary axoneme.

Recent evidence has revealed that the dynein motors and highly conserved signaling proteins are localized within the ciliary 9+2 axoneme. One key mechanism for regulation of motility is phosphorylation. Here, we review diverse evidence, from multiple experimental organisms, that ciliary motility is regulated by phosphorylation/dephosphorylation of the dynein arms through kinases and phosphatases that are anchored immediately adjacent to their axonemal substrates.