PROJECT SUMMARY
In the present proposal we will investigate new tissue processing protocols designed to preserve DNA, as well
as the morphology. We will preserve, process, and make available human ear tissue to inner ear basic
researchers they will be able to compare with their animal models. We have experience using high-resolution
advanced imaging techniques to the postmortem temporal bones, and also the membranous structures of the
inner ear. We propose to develop and apply these imaging techniques systematically and to correlate the
imaging findings with the temporal bone histopathology aiming to visualize structures with non-destructive
techniques. In Specific Aim 1) we will serve as part of a collaborative network providing technical services for
procuring, preparing, sectioning, and distributing high-quality human inner ear tissues aiming to increase
interest in the use of human inner ear tissues for basic scientists. We will continue to procure post- mortem
temporal bones aiming to minimize post-mortem times and will facilitate and reach out to additional prospective
researchers through the collaborative network and through meetings and provide human inner ear tissues of
high-quality. We will create hematoxylin and eosin digital shareable library with our archive of temporal bones,
and this library will be shared through the National Temporal Bone Registry to scientists. We will disseminate
protocols for temporal bone collection, processing, and inner ear tissue processing and provide hands-on
instruction on these protocols on demand and via zoom conferences. In Specific Aim 2, we will develop new
tissue collection and processing techniques for temporal bones by using animal temporal bones to develop
techniques which preserve DNA, mRNA and proteins and preserve morphology. We aim to develop new tissue
processing protocols which will allow for the use of immunofluorescence, DNA extraction and sequencing, and
the application of RNAScope, a powerful in situ hybridization tool. A new protocol will be developed to further
optimize sequencing the DNA from the extracted DNA using temporal bones specimens to correlate the gene
sequencing with the histopathology. In Aim 3. We will improve temporal bone inner ear visualization using
novel non-destructive imaging techniques which we will correlate with histopathology. In Aim 4, we will develop
a deep learning-based stain transformation framework to visualize specific anatomical structures such as the
spiral ganglion neurons to obtain accurate histopathological analysis.
