This study examined the effects of bilateral and unilateral deafness by measuring cortical auditory evoked potential (CAEP) responses in children at initial stages of bilateral cochlear implant (CI) use. We recorded cortical responses evoked by right and left CI stimulation in 127 children with early onset (< 12 months) deafness, with 72 children receiving the two devices in the same surgery (simultaneously implanted) and 55 children receiving the devices in separate procedures (sequentially implanted). Three different types of responses were identified in children with bilateral CIs. No significant effects of duration of deafness, age at implantation, or duration of unilateral CI use were found on response latencies and amplitudes within each type of cortical response, but there were clear differences in responses types between groups and ears. In the context of these findings, the effects of bilateral and unilateral deafness to the auditory pathways were discussed.

This study examined the effects of bilateral and unilateral deafness by measuring cortical auditory evoked potential (CAEP) responses in children at initial stages of bilateral cochlear implant (CI) use. We recorded cortical responses evoked by right and left CI stimulation in 127 children with early onset (< 12 months) deafness, with 72 children receiving the two devices in the same surgery (simultaneously implanted) and 55 children receiving the devices in separate procedures (sequentially implanted). Three different types of responses were identified in children with bilateral CIs. No significant effects of duration of deafness, age at implantation, or duration of unilateral CI use were found on response latencies and amplitudes within each type of cortical response, but there were clear differences in responses types between groups and ears. In the context of these findings, the effects of bilateral and unilateral deafness to the auditory pathways were discussed.

No description available.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19230号 / 医博第4029号 / 新制||医||1011(附属図書館) / 32229 / 京都大学大学院医学研究科医学専攻 / (主査)教授 渡邊 直樹, 教授 斎藤 通紀, 教授 村井 俊哉 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Microcephaly is a debilitating condition in which children are born with small brains. It can be caused by a variety of factors including maternal infection, harmful substance exposure, and genetic mutation. Cerebral cortical development is often severely disrupted in human microcephaly patients. In order to form the billions of neurons which exist in the cortex, efficient and correct neural progenitor division, differentiation, and migration are key. As the center of higher brain function in mammals, reduction in cortical mass is associated with the developmental delays that are symptomatic of microcephaly. Recently, a number of microcephaly causes have been linked to P53-mediated apoptosis of neural cells. The tumor suppressor protein P53 is upregulated in response to mitotic cycle stress, and its activation can trigger cell cycle arrest or apoptotic cell death. In microcephaly, P53 can become activated by mitotic stress and trigger apoptosis to cause the loss of cortical cell numbers that leads to microcephaly. Microcephaly has often been linked to mutations in mitotic proteins that alter neural progenitor division. However, the apical polarity complex Protein Associated with Lin-7 1 (PALS1) – known as membrane palmitoylated protein (MPP)5 in people – has recently been implicated in human microcephaly. PALS1 is integral to establishing polarity in neural progenitors. Deletion of Pals1 in mouse models has also resulted in microcephaly characterized by smaller brains and a global reduction in cortical cell numbers. Interestingly, a cellular phenomenon known as entosis can be caused by polarity disruptions in epithelial cells, and P53 activation has been shown to cause entosis in MDCK cell culture. While entosis is mainly associated with cancer cells, it is a form of competitive cell cannibalism that can eliminate unfit cells from a population. The loss of PALS1 from the developing cortex is known to result in apical polarity complex disruption and microcephaly in mouse models. However, the mechanism by which the loss of PALS1 results in cortical abrogation has yet to be determined. In Chapter 1 of this dissertation, I begin by reviewing cortical development. The normal progression of cortical cells from neural progenitors to fully differentiated neurons is explained in detail. Neural progenitor mitosis in particular is addressed in detail. Furthermore, an overview of microcephaly is provided to address the similarities between known causes of microcephaly. Next, I review the polarity complex proteins and their roles in cortical development. I compare and contrast the cortical phenotypes that have been described when each of the polarity complex proteins has been genetically deleted from the mouse cortex. I go on to review studies that have shown P53-mediated apoptosis in microcephaly in order to address the phenotypic features of microcephaly that are or are not caused by P53 activation. Finally, I provide a brief history of entosis. As a newly discovered cellular process in neural progenitors, the overview of entosis highlights what is known about cell cannibalism and the contexts in which it occurs. Following this background, I describe the experimental aims, hypotheses, and methods for this project in Chapter 2. In Chapter 3, I describe our investigation of three human patients with mutations in the Pals1 gene. One of the patients, possessing a heterozygous de novo nonsense mutation in Pals1 (or MPP5), was diagnosed with microcephaly. In order to model this patient’s phenotype, we generated a heterozygous conditional knockout of Pals1 from the entire mouse nervous system with Nestin-Cre. Through behavioral analysis of these mice, I demonstrate that they are hyperactive and blind, mimicking the microcephaly patient’s symptoms. Furthermore, via analysis of the mouse cortex, I show that heterozygous deletion of Pals1 results in severe microcephaly in mice with a global reduction in cortical cell numbers at both adult and embryonic stages. Importantly, I determine that Pals1 deletion does not result in proliferation or migration defects in the mouse cortex. Instead, loss of PALS1 results in massive apoptotic cell death that affects every cell type produced in the developing cortex. In Chapter 4, I detail our investigation into the mechanism underlying cell death in the PALS1-deficient cortex. By studying dividing neural progenitors at the apical surface in both Emx1-Cre and hGFAP-Cre drive Pals1 conditional knockout models, I demonstrate an as yet undescribed neural progenitor phenotype called entosis. As has been shown in cancer cells, neural progenitor entosis is dynamic and reliant on Rho-ROCK activity to occur. Furthermore, entosis produces observable cell-in-cell structures that persist through outer cell division and cause mitotic delay. I go on to demonstrate P53 activation in Pals1 deficient mouse cortices, and show that genetic deletion of Trp53 significantly rescues microcephaly. Trp53 deletion significantly restores all cortical cell types in addition to ameliorating entosis and mitotic length. This study suggests that P53 activation is a major mechanism by which PALS1 loss results in microcephaly. Overall, these studies show that deletion of Pals1 in mice can mimic microcephaly found in a human patient with a Pals1 mutation. Furthermore, PALS1 loss promotes P53-mediated cortical cell apoptosis. These studies provide the first description of entosis in neural progenitors, and suggest that entosis could be a mechanism for unfit cell removal in the developing cortex. Furthermore, I provide evidence that ROCK inhibition can fully rescue the presence of entosis in PALS1-deficient neural progenitors, and that genetic deletion of Trp53 significantly restores microcephaly pathology after PALS1 loss. These studies open up a field of research into the causes and effects of entosis in neural progenitors, and provide further evidence that apoptotic cell death in microcephaly is largely mediated by P53 activation. / Biomedical Sciences

Fundamental to mammalian brain development is the integration of cell intrinsic and extrinsic signals that direct the proliferation and differentiation of neural stem cells. Precise expression of transcription factors together with other intracellular components instruct progenitor cell fate, whereas interaction with extracellular signaling factors refines this process. We have elucidated the composition of the cerebrospinal fluid that is the source of multiple extrinsic cues during brain development. The choroid plexus, a highly vascularized tissue located in each ventricle of the brain, actively secretes cerebrospinal fluid. By RNA sequencing, we obtained transcriptome data on the choroid plexi from lateral and fourth ventricles of the mouse brain and discovered that they include transcripts unique to each tissue. Transcription factor expression in the macaque and human choroid plexi suggests that positional identities of these tissues are conserved in the primate brain. Based on transcriptional results, we defined the choroid plexus secretome, a prediction of secreted factors from the choroid plexus. By quantitative mass spectrometry, we detected proteins secreted by each choroid plexus, and comparison of these proteomic results with transcriptional profiling suggests that choroid plexus transcriptomes contribute to availability of regionalized cerebrospinal fluid factors during development. In the second part of my dissertation research, I studied the role of DNA repair mechanisms in regulating neural stem cells. These studies focused on DNA LigaseIV, an essential component of DNA double-stranded break repair, during cerebral cortical development. Deficiency of LigaseIV activity caused by a missense mutation leads to LigaseIV syndrome, in which a key clinical feature is microcephaly. Using the Lig4 R278H mouse mutant, we found increased cell death in the developing cortex, contributing to reduced cortical thickness and cellularity in the anterior cerebral cortex. These results indicate that DNA LigaseIV is essential for proper cortical development. Together, these findings illustrate the complexity of regulatory mechanisms that guide brain development, requiring the integration of mechanisms from within and outside the cell. We have investigated two such mechanisms, extrinsic cues from regionalized cerebrospinal fluid and DNA LigaseIV. These results should provide greater insight into mechanisms of normal brain development and neuropathological states. / 2017-11-02T00:00:00Z

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When it comes to cell types, the cerebral cortex is one of the most diverse regions in the mammalian brain. Mouse cortical neurons are generated during development from radial glial cells (RGCs). But how these stem cells generate the different neuronal subtypes is still an open question. In the adult, transcription factors, specially homeobox genes, have been identified as determinants of neuronal types throughout the animal kingdom. Thus, in this study, we hypothesise that different subpopulations of neuronal progenitors (RGCs) give rise to subsequent subtypes of neurons in the cortex, and these populations can be defined by homeobox gene expression. Starting from a scRNA- seq analysis, we identified differentially expressed genes across different progenitor populations in the developing cortex: Adnp2, Homez and Hmbox1. We characterised their mRNA and protein expression across cortical layers in postnatal mice and found that these genes are also differentially expressed among layers. We also find discordances between scRNA-seq data, mRNA expression, and protein expression data that could indicate specific post-transcriptional regulation of these genes. Altogether, these results point to a role of homeobox genes in neuronal identity.

No description available.