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High temperature requirement A serine peptidase 1 (HTRA1) is a serine protease involved in an array of signaling pathways. It is also responsible for the regulation of protein aggregates via refolding, translocation, and degradation. It has subsequently been found that runaway proteolytic HTRA1 activity plays a role in a variety of diseases, including Age-Related Macular Degeneration (AMD), osteoarthritis, and Rheumatoid Arthritis. Selective inhibition of serine protease HTRA1 therefore offers a promising new strategy for the treatment of these diseases. Herein we disclose structure-activity-relationship (SAR) studies which identify key interactions responsible for binding affinity of small molecule inhibitors to HTRA1. The study results in highly potent molecules with IC50's less than 15 nM and excellent selectivity following a screen of 35 proteases.
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Serina Peptidase 1 de Requerimento de Alta Temperatura A , Serina Endopeptidases , Serina Peptidase 1 de Requerimento de Alta Temperatura A/metabolismo , Relação Estrutura-Atividade , Humanos , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/farmacologia , Inibidores de Serina Proteinase/química , Inibidores de Serina Proteinase/síntese química , Estrutura Molecular , Relação Dose-Resposta a Droga , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/síntese químicaRESUMO
OBJECTIVES: Despite growing interest in the genetic contribution to cochlear implant (CI) outcomes, only a few studies with limited samples have examined the association of CI outcomes with genetic etiologies. We analyzed CI outcomes using known predictors and genetic testing results to obtain a comprehensive understanding of the impact of genetic etiologies. DESIGN: We retrospectively reviewed the medical records and images of patients who underwent cochlear implantation and genetic testing at a single tertiary medical institution, between May 2008 and December 2020. After excluding those whose speech test results were unavailable, and those in whom the implant was removed due to complications, such as infection or device failure, 203 patients were included in this study. The participants were categorized into adult (≥19 years), child (2-18 years), and infant (<24 months) groups. Outcomes were measured based on categories of auditory perception, monosyllable, disyllable, and sentence scores. For the infant group, the Infant-Toddler Meaningful Auditory Integration Scale score was used. RESULTS: Among the 203 participants, a causative genetic variant was identified in 117 (57.6%) individuals. The presence of a causative variant was significantly associated with better CI outcomes in the infant group (ß = 0.23; 95% confidence interval, 0.01 to 0.47; p = 0.044), but not in the child and adult groups. In the genetically confirmed patients without cochlear malformation, genetic variants involving the spiral ganglion was a poor prognostic factor in the child group (ß = -57.24; 95% confidence interval, -90.63 to -23.75; p = 0.004). CONCLUSIONS: The presence of known genetic etiology of hearing loss was associated with better CI outcomes in the infant group, but not in the child and adult groups. A neural-type genetic variant was a poor prognostic factor in the genetically diagnosed child subgroup without cochlear malformation. Careful genetic counseling should be performed before cochlear implantation.
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Implante Coclear , Implantes Cocleares , Percepção da Fala , Adulto , Lactente , Humanos , Implante Coclear/métodos , Estudos Retrospectivos , Resultado do Tratamento , Testes GenéticosRESUMO
BACKGROUND: Mutations in mitochondrial DNA (mtDNA) are associated with several genetic disorders, including sensorineural hearing loss. However, the prevalence of mtDNA mutations in a large cohort of Korean patients with hearing loss has not yet been investigated. Thus, this study aimed to investigate the frequency of mtDNA mutations in a cohort of with pre- or post-lingual hearing loss of varying severity. METHODS: A total of 711 Korean families involving 1,099 individuals were evaluated. Six mitochondrial variants associated with deafness (MTRNR1 m.1555A>G, MTTL1 m.3243A>G, MTCO1 m.7444G>A and m.7445A>G, and MTTS1 m.7471dupC and m.7511T>C) were screened using restriction fragment length polymorphism. The prevalence of the six variants was also analyzed in a large control dataset using whole-genome sequencing data from 4,534 Korean individuals with unknown hearing phenotype. RESULTS: Overall, 12 of the 711 (1.7%) patients with hearing loss had mtDNA variants, with 10 patients from independent families positive for the MTRNR1 m.1555A>G mutation and 2 patients positive for the MTCO1 m.7444G>A mutation. The clinical characteristics of patients with the mtDNA variants were characterized by post-lingual progressive hearing loss due to the m.1555A>G variant (9 of 472; 1.9%). In addition, 18/4,534 (0.4%) of the Korean population have mitochondrial variants associated with hearing loss, predominantly the m.1555A>G variant. CONCLUSION: A significant proportion of Korean patients with hearing loss is affected by the mtDNA variants, with the m.1555A>G variant being the most prevalent. These results clarify the genetic basis of hearing loss in the Korean population and emphasize the need for genetic testing for mtDNA variants.
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Perda Auditiva Neurossensorial , Perda Auditiva , Humanos , Prevalência , Perda Auditiva Neurossensorial/epidemiologia , Perda Auditiva Neurossensorial/genética , Mutação , DNA Mitocondrial/genética , República da Coreia/epidemiologiaRESUMO
This phenotype-genotype study aimed to investigate the extent of audioprofile variability related to cochlin major domains and to identify potential ethnic-specific differences associated with COCH-related hearing loss. Eight Korean families (26 cases) were diagnosed with COCH-related hearing loss by exome sequencing. Audiometric test results were combined with those from nine published East Asian families (20 cases) and compared with those from 38 European-descent families (277 cases). Audioprofiles were created by grouping audiometric test results into age ranges by age at testing and then averaging hearing loss thresholds by frequency within age ranges. The functional impact of the identified variants was assessed in vitro by examining the intracellular trafficking, secretion, and cleavage of cochlin. In both East Asian and European-descent families segregating COCH-related hearing loss, deafness-associated variants in non-LCCL domains of cochlin were associated with hearing loss that was more severe earlier in life than hearing loss caused by variants in the LCCL domain. Consistent with this phenotypic difference, functional studies demonstrated distinct pathogenic mechanisms for COCH variants in a domain-dependent manner; specifically, a cytotoxic effect was observed for the p.Phe230Leu variant, which is located in the vWFA1 domain. No ethnic-specific differences in hearing loss progression were observed, except for those attributable to an overrepresentation of presymptomatic cases in the European-descent cohort.
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Surdez , Perda Auditiva Neurossensorial , Perda Auditiva , Humanos , Surdez/genética , Proteínas da Matriz Extracelular/genética , Genótipo , Perda Auditiva/genética , Perda Auditiva Neurossensorial/genética , Mutação , Linhagem , FenótipoRESUMO
Ski-slope hearing loss (HL), which refers to increased auditory threshold at high frequencies, is common in adults. However, genetic contributions to this post-lingual HL remain largely unknown. Here, we prospectively investigated deafness-associated and novel candidate genes causing ski-slope HL. We analyzed 192 families with post-lingual HL via gene panel and/or exome sequencing. With an overall molecular diagnostic rate of 35.4% (68/192) in post-lingual HL, ski-slope HL showed a lower diagnostic rate (30.7%) compared with other conditions (40.7%). In patients who showed HL onset before the age of 40, genetic diagnostic probability was significantly lower for ski-slope HL than for other conditions. Further analysis of 51 genetically undiagnosed patients in the ski-slope HL group identified three variants in delta-like ligand 1 (DLL1), a Notch ligand, which presented in vitro gain-of-function effects on Notch downstream signaling. In conclusion, genetic diagnostic rates in post-lingual HL varied according to audiogram patterns with age-of-onset as a confounding factor. DLL1 was identified as a candidate gene causing ski-slope HL.
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Perda Auditiva Neurossensorial , Perda Auditiva , Adulto , Perda Auditiva/diagnóstico , Perda Auditiva/genética , Perda Auditiva Neurossensorial/genética , Testes Auditivos , Humanos , Ligantes , Patologia Molecular , Linhagem , Sequenciamento do ExomaRESUMO
Background: This study aimed to evaluate the injury mechanism of medial epicondylar fractures in children and adolescents and its association with increased carrying angle (CA) as a predisposing factor. Materials and Methods: We evaluated 37 patients with medial epicondylar fractures who were surgically treated at our institution. Medical records and plain radiographs were reviewed to determine the mechanism of injury and the humerus-elbow-wrist angle (HEWA) and CA of the uninjured arm. To evaluate the effect of coronal alignment on specific fracture type, we compared the CA and HEWA of the 23 patients with medial epicondylar fracture who were injured by falling onto an outstretched hand (group I) with age- and sex-matched controls of 23 patients who had sustained extension-type supracondylar fractures (group II). Results: The mean age at injury was 11.7 ± 2.8 years (range, 5 to 16 years). Of the 37 patients, 23 (62.2%) recalled the injury mechanism as falling onto an outstretched hand and 10 patients (27.0%) were injured while arm wrestling, and in one patient (2.7%), the injury was associated with elbow dislocation. In the case-matched analysis, the mean HEWA of group I was 13.1 ± 2.8° (range, 7.1° to 19.8°) and the mean CA was 17.7 ± 2.7° (range, 13.0° to 22.2°). These angles were significantly increased in group I (p=0.003 and p=0.001, respectively). Conclusion: Falling onto an outstretched hand is the most common injury mechanism in patients with medial epicondylar fractures, and these fractures are associated with an increased CA.
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Lesões no Cotovelo , Articulação do Cotovelo , Fraturas do Úmero , Adolescente , Criança , Articulação do Cotovelo/diagnóstico por imagem , Articulação do Cotovelo/cirurgia , Humanos , Fraturas do Úmero/diagnóstico por imagem , Fraturas do Úmero/cirurgia , Radiografia , Amplitude de Movimento Articular , Estudos Retrospectivos , Resultado do TratamentoRESUMO
Background: Although it is generally agreed that vitamin D is important for bone health, the role of vitamin D in preventing fractures in children and adolescents remains unclear. Therefore, this study aimed to investigate the prevalence of vitamin D deficiency and insufficiency in healthy Korean children with fractures. Our secondary aim was to compare serum vitamin D levels before and during the coronavirus disease 2019 (COVID-19) outbreak. Methods: We evaluated 334 patients with fractures who were surgically treated at our institution between 2018 and 2019 before the onset of COVID-19 (group I). In addition, we collected data on the serum 25(OH)D levels of 210 patients who visited our pediatric department for evaluation of short stature (group II) and the serum 25(OH)D levels of the patients with fractures during the COVID-19 pandemic period (group III). A serum 25(OH)D level of <20 ng/mL was considered deficient, between 20 and 32 ng/mL was insufficient, and ≥32 ng/mL was considered sufficient. Results: The mean age was 8.1 ± 3.5 years in group I, 8.2 ± 3.7 years in group II, and 8.6 ± 3.5 years in group III. The prevalence of vitamin D deficiency was 53.0% in group I and 32.9% in group II. The mean serum 25(OH)D level was lower in group I than in group II (20.0 ± 7.3 ng/ml vs. 23.2 ± 6.9 ng/ml, p < 0.001). The mean serum 25(OH)D level of younger patients (<10 years) in group III was lower than that of the younger patients in the prepandemic period (21.4 ± 7.2 ng/mL vs. 19.2 ± 6.8 ng/mL, p=0.037). Conclusions: We observed a high prevalence of vitamin D deficiency/insufficiency in children with fractures who required surgical treatment. During the COVID-19 pandemic, the serum vitamin D levels became even lower, especially in younger children.
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COVID-19 , Fraturas Ósseas , Deficiência de Vitamina D , Adolescente , COVID-19/complicações , COVID-19/epidemiologia , Criança , Pré-Escolar , Surtos de Doenças , Fraturas Ósseas/epidemiologia , Humanos , Pandemias , Prevalência , Vitamina D , Deficiência de Vitamina D/complicações , Deficiência de Vitamina D/epidemiologiaRESUMO
Mutations in potassium voltage-gated channel subfamily Q member 4 (KCNQ4) are etiologically linked to nonsyndromic hearing loss (NSHL), deafness nonsyndromic autosomal dominant 2 (DFNA2). To identify causative mutations of hearing loss in 98 Korean families, we performed whole exome sequencing. In four independent families with NSHL, we identified a cosegregating heterozygous missense mutation, c.140T>C (p.Leu47Pro), in KCNQ4. Individuals with the c.140T>C KCNQ4 mutation shared a haplotype flanking the mutated nucleotide, suggesting that this mutation may have arisen from a common ancestor in Korea. The mutant KCNQ4 protein could reach the plasma membrane and interact with wild-type (WT) KCNQ4, excluding a trafficking defect; however, it exhibited significantly decreased voltage-gated potassium channel activity and fast deactivation kinetics compared with WT KCNQ4. In addition, when co-expressed with WT KCNQ4, mutant KCNQ4 protein exerted a dominant-negative effect. Interestingly, the channel activity of the p.Leu47Pro KCNQ4 protein was rescued by the KCNQ activators MaxiPost and zinc pyrithione. The c.140T>C (p.Leu47Pro) mutation in KCNQ4 causes progressive NSHL; however, the defective channel activity of the mutant protein can be rescued using channel activators. Hence, in individuals with the c.140T>C mutation, NSHL is potentially treatable, or its progression may be delayed by KCNQ activators.
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Surdez/genética , Canais de Potássio KCNQ/genética , Mutação/genética , Adulto , Idoso , Animais , Células CHO , Pré-Escolar , Cricetinae , Cricetulus , Feminino , Células HEK293 , Humanos , Ativação do Canal Iônico , Cinética , Masculino , Pessoa de Meia-Idade , Linhagem , Fenótipo , Subunidades Proteicas/genética , República da Coreia , Sequenciamento do Exoma , Adulto JovemRESUMO
BACKGROUND: Trichophyton usually causes a superficial skin infection, affecting the outermost layer of the epidermis, the stratum corneum. In immunocompromised patients, deeper invasion into the dermis and even severe systemic infection with distant organ involvement can occur. Most cases of deeper dermal dermatophytosis described in the literature so far involved pre-existing superficial dermatophytosis. CASE PRESENTATION: We report a 68-year-old woman presented to our clinic with a 3-month history of palpable nodules on the right ankle without pre-existing superficial dermatophytosis. Magnetic resonance imaging showed multiple, well-demarcated, cystic lesions around the lateral malleolus, located in the subcutaneous or dermal layers. The sizes varied from 0.5 cm to 4 cm in diameter. The patient underwent complete excision of the lesions. Fungal culture yielded Trichophyton rubrum on Sabouraud dextrose agar. Histopathology showed organizing abscesses with degenerated fungal hyphae. After the 12-week oral itraconazole therapy, the lesions were completely resolved. CONCLUSION: Dermatophytes should be considered as a possible cause of deep soft tissue abscesses in immunocompromised patients, even though there is no superficial dermatophytosis lesion.
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Abscesso/microbiologia , Tinha/diagnóstico , Trichophyton/isolamento & purificação , Tornozelo , Antifúngicos/uso terapêutico , Feminino , Humanos , Itraconazol/uso terapêutico , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Real-time sonoelastography can be used to assess tissue elasticity. The present study evaluated the relationship between tendon stiffness on sonoelastography and the magnetic resonance imaging (MRI) tendinosis grade in patients with rotator cuff tendinopathy. METHODS: The study included 39 patients with chronic pain and no history of trauma or rotator cuff tear. The supraspinatus tendons were graded according to MRI findings (grade 0, normal; grade 1, mild tendinosis; grade 2, moderate tendinosis; grade 3, marked tendinosis), and the subcutaneous fat-to-tendon (Fat/T) and gel pad-to-tendon (Pad/T) strain ratios were assessed. We used the trend test to analyze the relationship of the MRI grade with the Fat/T strain ratio and the Pad/T strain ratio. RESULTS: Of the 39 patients, 9 had grade 0, 17 had grade 1, 12 had grade 2, and 1 had grade 3 tendinosis. The mean real-time elastography Fat/T and Pad/T strain ratios were 2.92 ± 2.13 and 20.77 ± 21.78 in patients with grade 0 tendinosis, 4.08 ± 4.09 and 21.78 ± 17.16 in patients with grade 1 tendinosis, 13.48 ± 10.19 and 83.00 ± 48.26 in patients with grade 2 tendinosis, and 12.3 ± 0.00 and 16.58 ± 0.00 in patients with grade 3 tendinosis, respectively. The Fat/T and Pad/T strain ratios were positively associated with the MRI grade (P <.001). CONCLUSION: The MRI tendinosis grade is associated with stiffness assessed using sonoelastography in patients with rotator cuff tendinopathy. Therefore, sonoelastography might be a useful diagnostic tool for supraspinatus tendinopathy. LEVEL OF EVIDENCE: Level III; Diagnostic Study.
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Técnicas de Imagem por Elasticidade , Imageamento por Ressonância Magnética , Manguito Rotador/diagnóstico por imagem , Tendinopatia/diagnóstico por imagem , Adulto , Dor Crônica/etiologia , Elasticidade , Técnicas de Imagem por Elasticidade/métodos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Manguito Rotador/fisiopatologia , Índice de Gravidade de Doença , Gordura Subcutânea/diagnóstico por imagem , Tendinopatia/complicaçõesRESUMO
PURPOSE: We present the clinical characteristics, radiographic, and bone scintigraphy finding of the cuboid fracture in early childhood. METHODS: From 2008 to 2012, we identified 25 patients (13 boys and 12 girls) with cuboid fracture who were seen in our institution. Medical records and radiographs as well as bone scintigraphy of 25 patients were reviewed. Nutcracker test was performed as a provocation test. RESULTS: The mean age of the patients was 24.7 months (range 15-38 months). The average duration of symptom before visit was 7 days (range 2-14 days). Most of the parents/caregivers (76 %) did not recall a traumatic episode. Patterns of limping were variable. Nutcracker test was positive in 11 patients. In 10 of 25 patients, initial plain radiographs of the foot showed no abnormal finding. The average duration of symptom of these patients was 4.5 days (range 2-7 days). In 15 patients, the radiograph of the foot showed sclerosis along the base of the cuboid. Bone scintigraphy of the patients with normal radiograph showed hot spot in cuboid. Eleven patients visited our institution to seek for second opinion as their child has persistent limping gait. CONCLUSIONS: It is not always possible to make an early diagnosis of this fracture since the initial radiographic finding and physical examination are often negative. Considering the consequences of a missed fracture and avoiding unnecessary treatment, bone scan might be useful in the early diagnosis of the stress fracture of the cuboid in young children. LEVEL OF EVIDENCE: Diagnostic study, Level IV.
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Fraturas de Estresse/diagnóstico por imagem , Ossos do Tarso/lesões , Pré-Escolar , Diagnóstico Precoce , Feminino , Marcha , Humanos , Lactente , Masculino , Radiografia , Cintilografia , Ossos do Tarso/diagnóstico por imagem , Suporte de CargaRESUMO
Introduction: Dysphagia is a common complication in patients with cervical spinal cord injury (C-SCI) and can cause various pulmonary complications, such as aspiration pneumonia and mechanical airway obstruction increasing mortality and morbidity. This study evaluated the clinical factors that predict dysphagia in patients with traumatic and non-traumatic C-SCI. Methods: Ninety-eight patients with C-SCI were retrospectively enrolled in this study and were divided into those with and without dysphagia. Clinical factors such as age, sex, tracheostomy, spinal cord independence measure, pulmonary function test (PFT) including forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1) and FVC/FEV1, American Spinal Cord Injury Association score, Berg Balance Scale, and surgical approach were investigated retrospectively. Results: Multivariate logistic regression analysis revealed that FVC and the presence of tracheostomy were significantly correlated with dysphagia in patients with C-SCI (p < 0.05). FVC and the presence of tracheostomy are useful tools for detecting dysphagia in patients with C-SCI. Conclusion: Considering the results of our study, early PFTs, especially FVC, in patients with C-SCI and early initiation of dysphagia management and treatment in patients with C-SCI and tracheostomy will be advantageous in lowering the mortality and morbidity due to pulmonary aspiration in these patients.
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KCNQ4 is a voltage-gated K+ channel was reported to distribute over the basolateral surface of type 1 vestibular hair cell and/or inner surface of calyx and heminode of the vestibular nerve connected to the type 1 vestibular hair cells of the inner ear. However, the precise localization of KCNQ4 is still controversial and little is known about the vestibular phenotypes caused by KCNQ4 dysfunction or the specific role of KCNQ4 in the vestibular organs. To investigate the role of KCNQ4 in the vestibular organ, 6-g hypergravity stimulation for 24 h, which represents excessive mechanical stimulation of the sensory epithelium, was applied to p.W277S Kcnq4 transgenic mice. KCNQ4 was detected on the inner surface of calyx of the vestibular afferent in transmission electron microscope images with immunogold labelling. Vestibular function decrease was more severe in the Kcnq4p.W277S/p.W277S mice than in the Kcnq4+/+ and Kcnq4+/p.W277S mice after the stimulation. The vestibular function loss was resulted from the loss of type 1 vestibular hair cells, which was possibly caused by increased depolarization duration. Retigabine, a KCNQ activator, prevented hypergravity-induced vestibular dysfunction and hair cell loss. Patients with KCNQ4 mutations also showed abnormal clinical vestibular function tests. These findings suggest that KCNQ4 plays an essential role in calyx and afferent of type 1 vestibular hair cell preserving vestibular function against excessive mechanical stimulation.
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Células Ciliadas Vestibulares , Canais de Potássio KCNQ , Camundongos Transgênicos , Animais , Canais de Potássio KCNQ/metabolismo , Canais de Potássio KCNQ/genética , Células Ciliadas Vestibulares/metabolismo , Células Ciliadas Vestibulares/patologia , Camundongos , Fenilenodiaminas/farmacologia , Carbamatos/farmacologia , Vestíbulo do Labirinto/metabolismo , Vestíbulo do Labirinto/patologia , Vestíbulo do Labirinto/fisiopatologiaRESUMO
Background: The vestibular phenotypes of patients with genetic hearing loss are poorly understood. Methods: we performed genetic testing including exome sequencing and vestibular function tests to investigate vestibular phenotypes and functions in patients with genetic hearing loss. Results: Among 627 patients, 143 (22.8%) had vestibular symptoms. Genetic variations were confirmed in 45 (31.5%) of the 143 patients. Nineteen deafness genes were linked with vestibular symptoms; the most frequent genes in autosomal dominant and recessive individuals were COCH and SLC26A4, respectively. Vestibular symptoms were mostly of the vertigo type, recurrent, and persisted for hours in the genetically confirmed and unconfirmed groups. Decreased vestibular function in the caloric test, video head impulse test, cervical vestibular-evoked myogenic potential, and ocular vestibular-evoked myogenic potential was observed in 42.0%, 16.3%, 57.8%, and 85.0% of the patients, respectively. The caloric test revealed a significantly higher incidence of abnormal results in autosomal recessive individuals than in autosomal dominant individuals (p = 0.011). The genes, including SLC26A4, COCH, KCNQ4, MYH9, NLRP3, EYA4, MYO7A, MYO15A, and MYH9, were heterogeneously associated with abnormalities in the vestibular function test. Conclusions: In conclusion, diverse vestibular symptoms are commonly concomitant with genetic hearing loss and are easily overlooked.
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Genetic hearing loss is the most common hereditary sensorial disorder. Though more than 120 genes associated with deafness have been identified, unveiled causative genes and variants of diverse types of hearing loss remain. Herein, we identified a novel nonsense homozygous variant in CEP250 (c.3511C>T; p.Gln1171Ter) among the family members with progressive moderate sensorineural hearing loss in nonsyndromic autosomal recessive type but without retinal degeneration. CEP250 encodes C-Nap1 protein belonging to the CEP protein family, comprising 30 proteins that play roles in centrosome aggregation and cell cycle progression. The nonsense variant in CEP250 led to the early truncating protein of C-Nap1, which hindered centrosome localization; heterologous expression of CEP250 (c.3511C>T) in NIH3T3 cells within cilia expression condition revealed that the truncating C-Nap1 (p.Gln1171Ter) was not localized at the centrosome but was dispersed in the cytosol. In the murine adult cochlea, Cep250 was expressed in the inner and outer hair cells. Knockout mice of Cep250 showed significant hair cell degeneration and progressive hearing loss in auditory brainstem response. In conclusion, a nonsense variant in CEP250 results in a deficit of centrosome localization and hair cell degeneration in the cochlea, which is associated with the progression of hearing loss in humans and mice.
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Pathogenic variants of KCNQ4 cause symmetrical, late-onset, progressive, high-frequency-affected hearing loss, which eventually involves all frequencies with age. To understand the contribution of KCNQ4 variants to hearing loss, we analyzed whole-exome and genome sequencing data from patients with hearing loss and individuals whose hearing phenotypes were unknown. In KCNQ4, we identified seven missense variants and one deletion variant in 9 hearing loss patients and 14 missense variants in the Korean population with an unknown hearing loss phenotype. The p.R420W and p.R447W variants were found in both cohorts. To investigate the effects of these variants on KCNQ4 function, we performed whole-cell patch clamping and examined their expression levels. Except for p.G435Afs*61, all KCNQ4 variants exhibited normal expression patterns similar to those of wild-type KCNQ4. The p.R331Q, p.R331W, p.G435Afs*61, and p.S691G variants, which were identified in patients with hearing loss, showed a potassium (K+) current density lower than or similar to that of p.L47P, a previously reported pathogenic variant. The p.S185W and p.R216H variants shifted the activation voltage to hyperpolarized voltages. The channel activity of the p.S185W, p.R216H, p.V672M, and p.S691G KCNQ4 proteins was rescued by the KCNQ activators retigabine or zinc pyrithione, whereas p.G435Afs*61 KCNQ4 proteins were partially rescued by sodium butyrate, a chemical chaperone. Additionally, the structure of the variants predicted using AlphaFold2 showed impaired pore configurations, as did the patch-clamp data. Our findings suggest that KCNQ4 variants may be overlooked in hearing loss that starts in adulthood. Some of these variants are medically treatable; hence, genetic screening for KCNQ4 is important.
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Surdez , Perda Auditiva , Humanos , Linhagem , Perda Auditiva/genética , Surdez/genética , Audição , Mutação de Sentido Incorreto , Canais de Potássio KCNQ/genéticaRESUMO
Autosomal dominant hearing loss (ADHL) manifests as an adult-onset disease or a progressive disease. MYO7A variants are associated with DFNA11, a subtype of ADHL. Here, we examined the role and genotype-phenotype correlation of MYO7A in ADHL. Enrolled families suspected of having post-lingual sensorineural hearing loss were selected for exome sequencing. Mutational alleles in MYO7A were identified according to ACMG guidelines. Segregation analysis was performed to examine whether pathogenic variants segregated with affected status of families. All identified pathogenic variants were evaluated for a phenotype-genotype correlation. MYO7A variants were detected in 4.7% of post-lingual families, and 12 of 14 families were multiplex. Five potentially pathogenic missense variants were identified. Fourteen variants causing autosomal dominant deafness were clustered in motor and MyTH4 domains of MYO7A protein. Missense variants in the motor domain caused late onset of hearing loss with ascending tendency. A severe audiological phenotype was apparent in individuals carrying tail domain variants. We report two new pathogenic variants responsible for DFNA11 in the Korean ADHL population. Dominant pathogenic variants of MYO7A occur frequently in motor and MyTH4 domains. Audiological differences among individuals correspond to specific domains which contain the variants. Therefore, appropriate rehabilitation is needed, particularly for patients with late-onset familial hearing loss.
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Intracellular accumulation of mutant proteins causes proteinopathies, which lack targeted therapies. Autosomal dominant hearing loss (DFNA67) is caused by frameshift mutations in OSBPL2. Here, we show that DFNA67 is a toxic proteinopathy. Mutant OSBPL2 accumulated intracellularly and bound to macroautophagy/autophagy proteins. Consequently, its accumulation led to defective endolysosomal homeostasis and impaired autophagy. Transgenic mice expressing mutant OSBPL2 exhibited hearing loss, but osbpl2 knockout mice or transgenic mice expressing wild-type OSBPL2 did not. Rapamycin decreased the accumulation of mutant OSBPL2 and partially rescued hearing loss in mice. Rapamycin also partially improved hearing loss and tinnitus in individuals with DFNA67. Our findings indicate that dysfunctional autophagy is caused by mutant proteins in DFNA67; hence, we recommend rapamycin for DFNA67 treatment.Abbreviations: ABR: auditory brainstem response; ACTB: actin beta; CTSD: cathepsin D; dB: decibel; DFNA67: deafness non-syndromic autosomal dominant 67; DPOAE: distortion product otoacoustic emission; fs: frameshift; GFP: green fluorescent protein; HsQ53R-TG: human p.Q53Rfs*100-transgenic: HEK 293: human embryonic kidney 293; HFD: high-fat diet; KO: knockout; LAMP1: lysosomal associated membrane protein 1; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; NSHL: non-syndromic hearing loss; OHC: outer hair cells; OSBPL2: oxysterol binding protein-like 2; SEM: scanning electron microscopy; SGN: spiral ganglion neuron; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; TG: transgenic; WES: whole-exome sequencing; YUHL: Yonsei University Hearing Loss; WT: wild-type.
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Surdez , Receptores de Esteroides , Animais , Humanos , Camundongos , Autofagia/genética , Surdez/genética , Células HEK293 , Camundongos Knockout , Camundongos Transgênicos , Proteínas Mutantes , Mutação/genética , Receptores de Esteroides/genética , Sirolimo/farmacologiaRESUMO
BACKGROUND: We investigated the reversibility of liver fibrosis induced with a CCl(4) injection and the role of stem cells in reversing the hepatic injury. Furthermore, the most effective cell fraction among bone marrow cells (BMCs) in the repair process was analysed. METHODS: C57BL/6 mice were divided into four groups after 5 weeks of injection of CCl(4) : control, sacrificed after 5 weeks, sacrificed at 10 weeks and sacrificed 5 weeks later after GFP-donor BM transplantation. Liver function tests and real-time polymerase chain reaction (PCR) of markers indicating liver fibrosis were compared between the groups. To identify the most effective BMC fraction that repairs liver injury, the mice were divided into three groups after the injection of CCl(4) for 2 days: granulocyte colony stimulating factor (G-CSF) only, mononuclear cell (MNC) transplantation and Lin-Sca-1+c-kit+haematopoietic stem cell (HSC) transplantation. Eight days after transplantation, the mice were harvested and morphometric, immunohistochemical analyses were performed to compare the expression of extracellular matrix and liver fibrosis-related factors. RESULTS: The liver fibrosis induced by CCl(4) was not spontaneously recovered but was persistent until 10 weeks, but the group injected with BMCs had less fibrosis and better liver function. Mobilization with G-CSF increased the recovery of the injured liver and the best results were seen in those mice administered the MNC fraction and Lin-Sca-1+c-kit+HSC fraction, with no difference between the two groups. CONCLUSION: BMC transplantation and stem cell mobilization with G-CSF effectively treats liver injury in mice. These are promising techniques for autologous transplantation in humans with liver fibrosis.
Assuntos
Transplante de Medula Óssea/métodos , Cirrose Hepática Experimental/terapia , Animais , Apoptose , Quimiocina CCL4/toxicidade , Primers do DNA/genética , Fator Estimulador de Colônias de Granulócitos/administração & dosagem , Transplante de Células-Tronco Hematopoéticas , Técnicas Histológicas , Imuno-Histoquímica , Leucócitos Mononucleares/transplante , Cirrose Hepática Experimental/induzido quimicamente , Testes de Função Hepática , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da PolimeraseRESUMO
Cell therapy using MSCs (mesenchymal stem cells) might be effective treatment for refractory GVHD (graft-versus-host disease). However, the fate and distribution of MSCs after transplantation remains unclear. In this study, an animal model was developed to monitor the dynamic distribution of MSCs in mice with GVHD. A GVHD mouse model was established by transplanting C57BL/6 donor bone marrow cells and C57BL/6 EGFP (enhanced green fluorescent protein) splenocytes into lethally irradiated BALB/c nude recipient mice. Donor MSCs were obtained from MHC-identical C57BL/6 RFP (red fluorescent protein) mice and infused into the recipient mice on the same transplantation day. In vivo movement of the donor splenocytes (EGFP) and MSCs (RFP) were evaluated by measuring the biofluorescence (IVIS-Xenogen system). Donor splenocytes and MSCs reached the lungs first, and then the gastrointestinal tract, lymph nodes and skin, in that order; the transit time and localization site of these cells were very similar. In the recipient mouse with GVHD, the number of detectable cells declined with time, as assessed by biofluorescence imaging and confirmed by RT (real-time)-PCR. This bioimaging system might be useful for preclinical testing and the design of therapeutic strategies for monitoring the dynamic distribution of MSCs with GVHD.