Evidence of progenitor cells in the adult human cochlea: sphere formation and identification of ABCG2

OBJECTIVES: The aim of this study was to search for evidence of stem or progenitor cells in the adult human cochlea by testing for sphere formation capacity and the presence of the stem cell marker ABCG2. METHODS: Cochleas removed from patients undergoing vestibular schwannoma resection (n=2) and from brain-dead organ donors (n=4) were dissociated for either flow cytometry analysis for the stem cell marker ABCG2 or a sphere formation assay that is widely used to test the sphere-forming capacity of cells from mouse inner ear tissue. RESULTS: Spheres were identified after 2-5 days in vitro, and the stem cell marker ABCG2 was detected using flow cytometric analysis after cochlear dissociation. CONCLUSIONS: Evidence suggests that there may be progenitor cells in the adult human cochlea, although further studies are required.

Desde la corteza auditiva a la cóclea: progresos en el sistema eferente auditivo / From auditory cortex to cochlea: progress in the auditory efferent system

El sistema eferente auditivo está constituido por el sistema olivococlear y por vías descendentes que provienen de la corteza auditiva y se dirigen a la cóclea. El sistema olivococlear se divide en una porción medial y una lateral, con neuronas que inervan a las células ciliadas externas y a fibras del nervio auditivo respectivamente. El principal neurotransmisor de las sinapsis olivococleares es acetilcolina, y tanto las células ciliadas externas como las fibras del nervio auditivo poseen receptores para esta molécula. El sistema eferente córtico-coclear se origina en la capa V y VI de la corteza auditiva y proyecta a los colículos inferiores y complejo olivar superior, donde a través del sistema olivococlear se conecta con el órgano receptor auditivo. En este artículo se revisan importantes hallazgos obtenidos en los últimos años que involucran (i) nuevos neurotransmisores y receptores del sistema eferente auditivo; (ii) vías descendentes de la corteza auditiva y su rol fisiológico sobre las respuestas cocleares y (iii) rol del sistema eferente auditivo en patologías audiológicas y neuropsiquiátricas.

The auditory efferent system is composed by the olivocochlear fibers and descending projections that originate in the auditory cortex and end in the cochlea. The olivocochlear system is divided into a medial and lateral division, with fibers directed to the outer hair cells and to the auditory nerve fibers respectively. It is known that acetylcholine is the main neurotransmitter of the olivocochlear synapses and that outer hair cells and auditory nerve fibers have receptors to this molecule. The cortico-cochlear efferent system originates in layers V and VI of the auditory cortex. These descending projections are directed to the inferior colliculus and superior olivary complex, a site in which the olivocochlear fibers emerge and connect the brain with the cochlear receptor. In this article recent discoveries obtained in the last years are reviewed: (i) new neurotransmitters and receptors of the olivocochlear system; (ii) anatomy and physiology of descending pathways from the auditory cortex to the cochlea and, (iii) clinical role of auditory efferents in audiological and neuropsychiatric pathologies.

Promoter effects of adeno-associated viral vector for transgene expression in the cochlea in vivo

The aims of this study were to evaluate the expression of enhanced green fluorescent protein (EGFP) driven by 6 different promoters, including cytomegalovirus IE enhancer and chicken beta-actin promoter (CAG), cytomegalovirus promoter (CMV), neuron-specific enolase promoter (NSE), myosin 7A promoter (Myo), elongation factor 1alpha promoter (EF-1alpha), and Rous sarcoma virus promoter (RSV), and assess the dose response of CAG promoter to transgene expression in the cochlea. Serotype 1 adeno-associated virus (AAV1) vectors with various constructs were transduced into the cochleae, and the level of EGFP expression was examined. We found the highest EGFP expression in the inner hair cells and other cochlear cells when CAG promoter was used. The CMV and NSE promoter drove the higher EGFP expression, but only a marginal activity was observed in EF-1alpha promoter driven constructs. RSV promoter failed to driven the EGFP expression. Myo promoter driven EGFP was exclusively expressed in the inner hair cells of the cochlea. When driven by CAG promoter, reporter gene expression was detected in inner hair cells at a dose as low as 3 x 10(7) genome copies, and continued to increase in a dose- dependent manner. Our data showed that individual promoter has different ability to drive reporter gene expression in the cochlear cells. Our results might provide important information with regard to the role of promoters in regulating transgene expression and for the proper design of vectors for gene expression and gene therapy.

Hair cell loss and regeneration in the chick cochlea after treatment with gentamicin

Hair cells can be damaged by countless agents, among them aminoglycoside antibiotics. In the cheek cochlea, the hair cell loss can be recovered by regeneration. The objectives of the present investigation were to estudy the time progression of injury caused by gentamicin and the regeneration process in chick hair cells. Gentamicin was administered in a single subcutaneous dose of 125 or 250 mg/kg to two groups of 3-day old chicks. The cochleae were processed for analysis by scanning electron microscopy on the 1st, 3rd, 5th and 20th day after injection The cellular sequence of degeneration and regeneration was studied. On the 20th day, most it he damaged cochlear area showed regenerated hair and suport cells. Stereocilia and microvilli were observed on the apical surface of the regenerated hair cells.

Localización histoquímica e inmunohistoquímica de fibrocitos en la cóclea de la rata / Histochemical and immunohistochemical location of fibrocyts in the cochlea of rats

El fibrocito, célula de origen mesenquimal del ligamento espiral de la coclea, ha sido investigado usando tecnicas inmunohistoquimicas que al intentar reproducirlas fallan debido a la impredecible alteracion de sitios antigenicos introducidos durante el procesamiento de la muestra. El presente estudio describe la estandarizacion de una tecnica histoquimica e inmunohistoquimica para la localizacion del fibrocito en la coclea de la rata. Dos ejemplares, bajo anestesia general con nembutal recibieron una inyeccion intracardiaca de fijadores tipo aldehidos, buffer salino fosfato y sales precipitantes. Luego de la muerte del animal su cabeza se sometio a postfijacion y decalcificacion, para realizarle cortes con microtomo cada 5 micrometros y tincion cada decima placa con hematoxilina-eosina. Finalmente, se utilizó inmunohistoquimica indirecta con el metodo Avidina-biotina conjugada y marcador monoclonal anti-vimentina y la proteina S100, lográndose la identificacion de los fibrocitos y demás componentes del organo de corti murino. La estandarizacion de esta tecnica es un hecho trascendental para el proceso de investigacion histopatologica del oido interno, aplicable al estudio de patologias que afecten a los fibrocitos del ligamento espiral

Células ciliadas isoladas em cultura: um novo modelo para estudo da ototoxicidade / Ciliated cells isolated in culture: a new model for the study of otoxicity

A otoxicidade da Amicacina e a proteçäo por gangliosídeos das células ciliadas externas (CCE) cocleares de cobaias foram estudadas com 3 grupos de 8 animais. O grupo I, controle, o grupo II recebeu Amicacina e o grupo III Amicacina e gangliosídeo. utilizando-se o método de isolamento mecânico das CCE em cultura, 179 +-33 células em cada cobaia normal foram encontradas conservando suas características vitais até 2 horas e meia. O padräo de degeneraçäo vital foi observado e descrito. As lesöes provocadas pela Amicacina foram intensas. A análise estatística foi significativa na comparaçäo entre o grupo I e os grupos II e III, mas o resultado entre os grupos II e III näo demonstraram efeitos significativos na proteçäo das células pelos gangliosídeos contra os efeitos da Amicacina. Concluiu-se que esta técnica utilizada pela primeira vez para estudo da otoxicidade, constitui um modelo válido importante, simples e rápido, que permite estudo minucioso das lesöes em células isoladas "in vitro" provocadas por drogas ototóxicas