2019

Integrative Systems Biology of Hearing Restoration

Integrative Systems Biology of Hearing Restoration
Seth Ament, Ph.D., University of Maryland School of Medicine

This project will focus on integrating multiple datasets from the HRP to gain insight into hair cell development and regeneration and prioritize specific "driver" genes that can be targeted to induce regeneration. The main premise is that we will be able to regenerate hair cells if we activate the correct set of hair cell–promoting genes in supporting cells. This process is called transdifferentiation, and it occurs naturally in species such as birds and fish, but not in the inner ear of adult mammals. HRP researchers have generated numerous genomic datasets that describe cochlear development and transdifferentiation in multiple species. By analyzing all of these data together using sophisticated network analysis tools, we aim to identify which genes are involved in these processes as well as key differences that may explain the inability of human and mouse cells to transdifferentiate. Finally, this will enable the identification of genes that can be targeted to enable transdifferentiation.

Comparison of Three Reprogramming Cocktails in the Organ of Corti: Cells, Transcriptomes, and Epigenomes

Comparison of Three Reprogramming Cocktails in the Organ of Corti: Cells, Transcriptomes, and Epigenomes
Andy Groves, Ph.D., Baylor College of Medicine

In the past year we have been investigating whether we are able to use genetic reprogramming techniques to generate new hair cells in the mouse cochlea. The results of this work are extremely promising: We were able to turn nonsensory cells of the mouse cochlea into hair cells. However, we consistently find that supporting cells in our mouse models do not respond to reprogramming. This is curious, as supporting cells are the cells responsible for producing new sensory hair cells in birds and fish. In the coming year, we will first examine supporting cells after our reprogramming attempts to see whether they have been able to activate any aspects of a hair cell program. Second, we will test whether hair cell death alters the adjacent supporting cells so that they become more responsive to reprogramming.

Detection of Transcriptome Changes in Single Cells After Aminoglycoside-Induced Hair Cell Loss in the Chicken Basilar Papilla

Detection of Transcriptome Changes in Single Cells After Aminoglycoside-Induced Hair Cell Loss in the Chicken Basilar Papilla
Stefan Heller, Ph.D., Stanford University

Birds regenerate cochlear hair cells by activating dormant supporting cells. This project builds on innovative methods and findings to study how supporting cells are activated when ototoxic drugs cause hair cell death. The project uses single cell analysis (during which we study the complement of genes that are active in many individual cells) to identify triggers that initiate, execute, sustain, and ultimately terminate the regenerative process. Using bioinformatics methods to process the resulting data, we will focus this year on the analysis of the chicken cochlea cell populations isolated at various time points during hair cell regeneration, which will reveal the molecular steps that occur during hair cell regeneration. We have already identified a first candidate gene signaling pathway that may regulate the regenerative process in the chicken cochlea, and we will be confirming that this pathway plays a role in hair cell regeneration. Interestingly, this pathway is not active during inner ear development, which sets it apart from other pathways linked to chicken (and zebrafish) hair cell regeneration; the other pathways are involved in cell development and may not represent a unique regenerative trigger. Finding triggers that specifically control regeneration may be an important stepping stone on the path to developing cures for hair cell loss in mice and, eventually, humans.

Epigenetics Analysis of Maturation and Regenerative Responses in the Mouse Organ of Corti and Utricle (2019)

Epigenetics Analysis of Maturation and Regenerative Responses in the Mouse Organ of Corti and Utricle
Neil Segil, Ph.D., University of Southern California

Although hair cell regeneration does not occur in mammals, newborn mice harbor a latent capacity for some regenerative responses. However, this capability disappears within the first few weeks of life. This observation provides an experimental window that this proposal exploits to address fundamental questions about the failure of hair cell regeneration in mammals. Specifically, we propose experiments to identify those changes in the genetic material, the chromatin, that are responsible for orchestrating the differentiation of new hair cells within the newborn organ of Corti in the inner ear, and to investigate the changes in the chromatin that lead to the failure of regeneration in the adult mammalian inner ear.

Implementing the gEAR for Data Sharing Within the HRP

Implementing the gEAR for Data Sharing Within the HRP
Ronna Hertzano, M.D., Ph.D., University of Maryland School of Medicine

When a group of geographically dispersed scientists collaborate on hair cell regeneration in three different animal models—chicken, zebrafish, and mouse—and use multiple methods to track how genes “instruct” cells (multi-omics), an enormous amount of data results. The work of visualizing, conceptualizing, and analyzing these data presents a considerable challenge, and as technology has advanced, much of the multi-omic data is generated at the single cell level, resulting in datasets and files that are too big to process with traditional tools, such as Excel worksheets. The gEAR portal (gene Expression Analysis Resource, umgear.org) responds to this need by enabling meaningful visualization and analysis of these complex datasets in the public or private domain—no advanced programming skills required. It has also evolved to become a primary data sharing, visualization, and analysis tool for auditory researchers outside of the HRP to become a platform that supports the hearing research community at large.

Mouse Model Systems to Interrogate Candidate Genes for Sensory Hair Cell Regeneration

Mouse Model Systems to Interrogate Candidate Genes for Sensory Hair Cell Regeneration
John Brigande, Ph.D., Oregon Health & Science University

Given what the HRP has discovered thus far, this project proceeds from the assumption that manipulation of multiple genes that regulate diverse signaling pathways may be required to reprogram supporting cells to become functional hair cells. We must first verify the expression pattern of candidate genes in the inner ear, then develop methods that allow us to modulate multiple candidate genes in each supporting cell in order to transmit the genetic instructions that trigger regeneration. This proposal’s first goal is to establish improved genome editing via oviductal nucleic acids delivery (iGONAD) technology, allowing us to identify as well as eliminate candidate genes. Our second goal is to turn on multiple genes simultaneously in the same cell using a mouse model. The overall objective is to establish a rapid method to describe candidate gene expression and function in the inner ear at any stage, and to define an approach to determine which genes are critical to modulate signaling pathways for hair cell regeneration.

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