[Mutation analysis and prenatal diagnosis for 12 families affected with hereditary hearing loss and enlarged vestibular aqueduct].

OBJECTIVE: To carry out mutation analysis and prenatal diagnosis for 12 families affected with hearing loss and enlarged vestibular aqueduct from southern Zhejiang province. METHODS: Clinical data and peripheral venous blood samples of 38 members from the 12 families were obtained. Mutations of 4 genes, namely SLC26A4, GJB2, c.538C to T and c.547G to A of GJB3, m.1555A to G and m.1494C to T of 12S rRNA, were detected by PCR and Sanger sequencing. Maternal contamination was excluded by application of STR detection during prenatal diagnosis. RESULTS: Among the probands from the 12 families, 11 were found to be compound heterozygotes or homozygotes and 25 were heterozygotes. All of the families were detected with IVS7-2A to G mutations, and 4 had a second heterozygous mutation (c.2168A to G of the SLC26A4 gene). Four rare pathogenic mutations, namely IVS5-1G to A, c.946G to T, c.1607A to G and c.2167C to G, were detected in another four families. In addition, the partner of proband from pedigree 3 was identified with compound heterozygous mutations of c.235delC and c.299-300delAT, and proband of pedigree 5 has carried a mutation of c.109G to A in GJB2. For SLC26A4 gene, prenatal diagnostic testing has revealed heterozygous mutations in 6 fetuses and compound heterozygous mutations in 2 fetuses. CONCLUSION: IVS7-2A to G and c.2168A to G of the SLC26A4 gene were the most common mutations in southern Zhejiang. Such mutations can be found in most families affected with hearing loss and enlarged vestibular aqueduct, which may facilitate genetic counseling and prenatal diagnosis for such families.

Mouse model of enlarged vestibular aqueducts defines temporal requirement of Slc26a4 expression for hearing acquisition.

Mutations in human SLC26A4 are a common cause of hearing loss associated with enlarged vestibular aqueducts (EVA). SLC26A4 encodes pendrin, an anion-base exchanger expressed in inner ear epithelial cells that secretes HCO3- into endolymph. Studies of Slc26a4-null mice indicate that pendrin is essential for inner ear development, but have not revealed whether pendrin is specifically necessary for homeostasis. Slc26a4-null mice are profoundly deaf, with severe inner ear malformations and degenerative changes that do not model the less severe human phenotype. Here, we describe studies in which we generated a binary transgenic mouse line in which Slc26a4 expression could be induced with doxycycline. The transgenes were crossed onto the Slc26a4-null background so that all functional pendrin was derived from the transgenes. Varying the temporal expression of Slc26a4 revealed that E16.5 to P2 was the critical interval in which pendrin was required for acquisition of normal hearing. Lack of pendrin during this period led to endolymphatic acidification, loss of the endocochlear potential, and failure to acquire normal hearing. Doxycycline initiation at E18.5 or discontinuation at E17.5 resulted in partial hearing loss approximating the human EVA auditory phenotype. These data collectively provide mechanistic insight into hearing loss caused by SLC26A4 mutations and establish a model for further studies of EVA-associated hearing loss.

New insight into the bony labyrinth: a microcomputed tomography study.

OBJECTIVES: To visualize and quantify the morphology and mineralization of the developing fetal human bony labyrinth, using 3D-microcomputed tomography (3D-microCT) imaging. METHODS: Eleven right temporal bones from late second and third trimester fetuses were used in this prospective pilot study. After fixation in 10% formalin solution, all samples underwent a microcomputed tomography (microCT) scan, permitting the 3D imaging of the bony labyrinth as well as the quantitative assessment of mineral density, angular distances and dimensions of inner ear components the progression of ossification was precised with histological observations. RESULTS: Our findings show different rates of growth among the semicircular canals, the vestibular aqueduct, the oval window, the round window and the cochlea. The final sizes of the cochlea and round window are achieved at 23 weeks of gestation, with heights of 5mm and 2mm, respectively. The oval window reaches adult size at 35 weeks, whereas the vestibular aqueduct will attain adult size after birth. An increasing degree of torsion of each semicircular canal is observed during fetal development. The superior semicircular canal achieves adult size at 24 weeks, before the posterior and the lateral canals (25 weeks). The time-course of ossification and mineralization observed in structures and confirmed by histology. CONCLUSIONS: During this developmental period poorly studied until now, our findings suggest that each part of the bony labyrinth follows distinct growth and ossification kinetics trajectories, some of these reaching their adult size only after birth.

Lack of pendrin expression leads to deafness and expansion of the endolymphatic compartment in inner ears of Foxi1 null mutant mice.

Mice that lack the winged helix/forkhead gene Foxi1 (also known as Fkh10) are deaf and display shaker/waltzer behavior, an indication of disturbed balance. While Foxi1 is expressed in the entire otic vesicle at E9.5, it becomes gradually restricted to the endolymphatic duct/sac epithelium and at E16.5 Foxi1 expression in the inner ear is confined to this epithelium. Histological sections, paintfill experiments and whole-mount hybridizations reveal no abnormality in inner ear development of Foxi1(-/-) mice before E13.5. Between E13.5 and E16.5 the membranous labyrinth of inner ears from null mutants starts to expand as can be seen in histological sections, paint-fill experiments and three-dimensional reconstruction. Postnatally, inner ears of Foxi1(-/-) mice are extremely expanded, and large irregular cavities, compressing the cerebellum and the otherwise normal middle ear, have replaced the delicate compartments of the wild-type inner ear. This phenotype resembles that of the human sensorineural deafness syndrome Pendred syndrome, caused by mutations in the PDS gene. In situ hybridization of Foxi1(-/-) endolymphatic duct/sac epithelium shows a complete lack of the transcript encoding the chloride/iodide transporter pendrin. Based on this, we would like to suggest that Foxi1 is an upstream regulator of pendrin and that the phenotype seen in Foxi1 null mice is, at least in part, due to defective pendrin-mediated chloride ion resorption in the endolymphatic duct/sac epithelium. We show that this regulation could be mediated by absence of a specific endolymphatic cell type--FORE (forkhead related) cells--expressing Foxi1, Pds, Coch and Jag1. Thus, mutations in FOXI1 could prove to cause a Pendred syndrome-like human deafness.

[Analysis of growth model in vestibular aqueduct during human fetal development].

OBJECTIVE: To characterize the growth of the developing vestibular aqueduct in order to gain a better understanding of the possible origins of enlarged vestibular aqueduct. METHODS: Serial sections from 32 temporal bones for every other week from human embryos ranging in age from 6 to 38 weeks were studied with 3DMed medical image analysis software. The internal, external aperture, midpoint diameter and length vestibular aqueduct were analyzed with regression analysis to obtain a growth curve. RESULTS: The internal aperture of vestibular aqueduct was began to form at 6 weeks' gestation. It grew to reach the posterior surface of the petrous bone in the posterior cranial fossa by 10 weeks' gestation. All width parameter mean value in our embryos nerve reached the maximum average width in the adult. Statistical analysis showed that the vestibular aqueduct grew in a nonlinear continuous fashion and instability trend throughout embryonic life, except length parameter. CONCLUSION: The vestibular aqueduct grows in a nonlinear fashion throughout embryonic life. The widest aqueduct measured in embryonic life does not reach the maximum average width in the adult. These results suggest that it would be possible for vestibular aqueduct to develop postnatally.

Embryological development and large vestibular aqueduct syndrome.

OBJECTIVES/HYPOTHESIS: Large vestibular aqueduct syndrome (LVAS) is a significant cause of hearing loss in early childhood. Many theories on the origins and causes of LVAS have been proposed, including arrest or maldevelopment of the vestibular labyrinth in embryonic life. Prior studies have described postnatal and adult vestibular aqueduct anatomy, but none has analyzed aqueduct growth throughout embryonic life. This study was undertaken to characterize the growth of the developing vestibular aqueduct to gain a better understanding of the possible origins of LVAS. STUDY DESIGN: Basic science, temporal bone histopathological study. METHODS: Serial sections from 48 temporal bones from human embryos ranging in age from 5 weeks' gestation to full term were studied with computer image analysis. Measurements of vestibular aqueduct internal and external aperture, midportion diameter, and length were analyzed to obtain a growth model of development. RESULTS: The vestibular aqueduct grows in a nonlinear fashion throughout embryonic life. All parameters fit a similar growth curve and never reached a maximum or began narrowing during development. Growth in one parameter correlated well with growth of another. There was good side-to-side correlation with all but the external aperture. CONCLUSIONS: Most of the membranous labyrinth reaches adult size by 20 weeks' gestation, but the vestibular aqueduct grows throughout embryonic life. The measurements and growth model obtained in this study are not consistent with the theory that LVAS results from an arrest in development early in fetal life. The data suggest that LVAS may result from postnatal and early childhood maldevelopment.

Imaging findings of the developing temporal bone in fetal specimens.

PURPOSE: To trace the development of the normal fetal temporal bone by means of plain radiography, MR, and CT. METHODS: Eighteen formalin-fixed fetal specimens, 13.5 to 24.4 weeks' gestational age, were examined with a mammographic plain film technique, CT, and MR imaging at 1.5 T. Temporal bone development and ossification were assessed. RESULTS: The membranous labyrinth grows with amazing rapidity and attains adult size by the middle of the gestation period. The cochlea, vestibule, and semicircular canals are very prominent and easily recognized on MR images. The otic capsule develops from a cartilage model. Ossification of the otic capsule proceeds rapidly between 18 and 24 weeks from multiple ossification centers that replace the cartilaginous framework. The mastoid, internal auditory canal, vestibular aqueduct, and external auditory canal continue to grow after birth. CONCLUSION: The study of fetal developmental anatomy may lead to a better understanding of congenital disorders of the ear. Faster MR scanning techniques may provide a method for in utero evaluation of the fetal temporal bone.

Unilateral duplication of the vestibular aqueduct.

A case of unilateral duplication of the vestibular aqueduct is presented. This very rare abnormality can be explained either by an excessive induction by the rhombencephalon or by an abnormal complete division of the endolymphatic duct into utricular and saccular parts.