Olfactory epithelium as an infinitive source of neural stem cells for derivation of inner ear hair cells / El epitelio olfatorio como fuente infinita de células madre neurales para la derivación de los vestibulocitos del oído interno

Surgical techniques for treatment of sensory neural hearing loss (SNHL) have unpredictable outcomes and in recent years cell therapy investigated for treatment of SNHL. Olfactory epithelium proceed neurogenesis during life time and provide an easily accessible source of neural stem cells. So the aim of this study was isolating neural stem cells from olfactory epithelium of rat and differentiation of these cells into hair cells of inner ear in vitro. The epithelium tissue of olfactory mucosa of rats were removed and digested by collagenase H. The digested tissue was cultured in flasks in suspension forms to create spheres. Spheres were passaged and from passage 2 spheres selected for differentiation. At this stage cells of spheres isolated from each other and placed in flask containing defined differentiation medium. Cells at this stage cultured in adhesive form. Immunohistochemistry and RT-PCR were used for neural stem cells and hair cells identification. Spheres formed from olfactory epithelium culture and immunohistochemistry revealed that cells of spheres from passage one and two expressed the neural stem cells markers. After culture of isolated cells in differentiation medium, the morphology of cells begun to change. The cells presented neural cells projections and after 10 days the projections elongated more and interact to each other in multi layers. RT-PCR and immunohistochemistry revealed that differentiated cells expressed hair cells specific genes. In this study we showed that neural stem cells of olfactory epithelium can differentiate into hair cells of inner ear and therefore can be used for treatment of SNHL.

Las técnicas quirúrgicas para el tratamiento de la pérdida auditiva neural sensorial (PANS) tienen resultados impredecibles y en los últimos años la terapia celular ha sido investigada para su tratamiento. El epitelio olfatorio se forma durante la neurogénesis y proporciona una fuente fácilmente accesible de células madre neurales. El objetivo de este estudio fue aislar las células madre neurales del epitelio olfativo de la rata y la diferenciación de estas células en vestibulocitos del oído interno in vitro. Se retiró el tejido del epitelio de la mucosa olfatoria de ratas y fue digerido con colagenasa H. El tejido se cultivó en forma de suspensión para crear esferas. Se seleccionaron dos esferas para la diferenciación. En esta fase, las células de esferas fueron aisladas unas de otras y colocadas en un medio de diferenciación definido. Células en esta etapa fueron cultivadas en forma adhesiva. Inmunohistoquímica y RT-PCR se utilizó para las células madre neurales y la identificación de células ciliadas. Las esferas formadas a partir del cultivo del epitelio olfatorio y la inmunohistoquímica revelaron que las células de esferas en etapas uno y dos expresaban los marcadores de células madre neurales. Se observaron cambios en la morfología de las células después del cultivo de células aisladas. RT-PCR e inmunohistoquímica revelaron que las células diferenciadas expresaron células específicas de gen de vestibulocitos. Se observó que las células madre neuronales de epitelio olfatorio puede diferenciarse en células en forma de cabello del oído interno y por lo tanto puede ser utilizado para el tratamiento de PANS.

What do we know about the neurogenic potential of different stem cell types? / O que sabemos sobre o potencial neurogênico de diferentes tipos de células-tronco?

Cell therapies, based on transplantation of immature cells, are being considered as a promising tool in the treatment of neurological disorders. Many efforts are being concentrated on the development of safe and effective stem cell lines. Nevertheless, the neurogenic potential of some cell lines, i.e., the ability to generate mature neurons either in vitro or in vivo, is largely unknown. Recent evidence indicate that this potential might be distinct among different cell lines, therefore limiting their broad use as replacement cells in the central nervous system. Here, we have reviewed the latest advancements regarding the electrophysiological maturation of stem cells, focusing our attention on fetal-derived-, embryonic-, and induced pluripotent stem cells. In summary, a large body of evidence supports the biological safety, high neurogenic potential, and in some diseases probable clinical efficiency related to fetal-derived cells. By contrast, reliable data regarding embryonic and induced pluripotent stem cells are still missing.

Terapias celulares, baseadas no transplante de células imaturas, têm sido consideradas ferramentas promissoras no tratamento de doenças neurológicas. Muitos esforços têm sido concentrados no desenvolvimento de linhas de células-tronco seguras e eficazes. No entanto, o potencial neurogênico de algumas linhagens celulares, ou seja, a habilidade de gerar neurônios maduros, in vitro ou in vivo, ainda é altamente desconhecida. Dados recentes sugerem que esse potencial é distinto entre diversos tipos celulares, o que limitaria o largo emprego como células restauradoras no sistema nervoso central. Neste relato, revisaram-se os avanços recentes relacionados à maturação eletrofisiológica de células-tronco, com foco em células derivadas de tecido fetal, células embrionárias e células pluripotentes induzidas. Em resumo, há evidências que apontam para segurança biológica de células fetais, com alto potencial neurogênico e, em se tratando de algumas doenças, provável eficiência clínica. Ao contrário, ainda não há dados confiáveis acerca de células embrionárias e pluripotentes induzidas.

A cell junction pathology of neural stem cells leads to abnormal neurogenesis and hydrocephalus

Most cells of the developing mammalian brain derive from the ventricular (VZ) and the subventricular (SVZ) zones. The VZ is formed by the multipotent radial glia/neural stem cells (NSCs) while the SVZ harbors the rapidly proliferative neural precursor cells (NPCs). Evidence from human and animal models indicates that the common history of hydrocephalus and brain maldevelopment starts early in embryonic life with disruption of the VZ and SVZ. We propose that a "cell junction pathology" involving adherent and gap junctions is a final common outcome of a wide range of gene mutations resulting in proteins abnormally expressed by the VZ cells undergoing disruption. Disruption of the VZ during fetal development implies the loss of NSCs whereas VZ disruption during the perinatal period implies the loss of ependyma. The process of disruption occurs in specific regions of the ventricular system and at specific stages of brain development. This explains why only certain brain structures have an abnormal development, which in turn results in a specific neurological impairment of the newborn. Disruption of the VZ of the Sylvian aqueduct (SA) leads to aqueductal stenosis and hydrocephalus, while disruption of the VZ of telencephalon impairs neurogenesis. We are currently investigating whether grafting of NSCs/neurospheres from normal rats into the CSF of hydrocephalic mutants helps to diminish/repair the outcomes of VZ disruption.