<p>Proper development of the inner ear, including the cochlea,
is necessary for normal hearing.
Development of the inner ear requires many signaling molecules under
both spatial and temporal control. These
signaling molecules include the wingless-related integration site (Wnts) and
the fibroblast growth factors (Fgfs) gene families. The embryonic chick inner ear was chosen as
the model to study cochlear development due to its homology with the mammalian
cochlea and the ease of access to the inner ear in ovo. Both the mammalian cochlea and the homologous
chick basilar papilla contain two domains with their own type of hair cells and
innervation. The neural side of the
basilar papilla contains the tall hair cells innervated by the afferent axons
which takes the noise signal to the brain.
The abneural side of the basilar papilla contains the short hair cells
innervated by the efferent axons which receive signals from the brain to turn
down added gain.</p>
<p>Previous research showed that virally induced cWnt9a
overexpression within the basilar papilla generated a neural side phenotype
across the basilar papilla (Munnamalai et al., 2017). These basilar papillas contained more tall
hair cells and increased innervation at embryonic day 18 (E18) than their
wild-type counterparts. Additionally,
there were many differentially expressed genes found to be downstream of cWnt9a
including cFgf3 and cFgf19. This project
focused on determining the role of cFgf19 in inducing a neural side phenotype
in the basilar papilla. First, in situ
hybridization was used to determine the cFgf3 and cFgf19 mRNA transcript
location with cWnt9a overexpression.
Both Fgfs were found across the basilar papilla. Next, a possible cWnt9a receptor, cFzd4,
which was upregulated with cWnt9a overexpression, was found in the neural side
of the basilar papilla. cFgf19 was then
overexpressed using one of two different vectors: RCAS(A)/EGFP-P2A-Fgf19 or
RCAS(B)/Fgf19-P2A-EGFP in which the order of cFgf19 transcription was
altered. RCAS(B)/Fgf19-P2A-EGFP was
found to produce less GFP when transfected into DF-1 cells than
RCAS(A)/EGFP-P2A-Fgf19. Additionally,
RCAS(B)/Fgf19-P2A-EGFP transfected cells produced secreted fusion proteins of
GFP and Fgf19, compared to RCAS(A)/EGFP-P2A-Fgf19 transfected cells which
produced secreted individual proteins.
The viruses were injected into the otocyst at E3 and the embryos
harvested several days later including at E6, E10, and E14. Inner ears injected with either virus showed
no changes in innervation, hair cells, proliferation, cartilage formation
around the cochlear duct, cFgf3 expression, or phosphorylation of ERK. To determine understand where Fgf19 could be
producing an effect, the location of a possible receptor, Fgfr4, was determined
in wild-type embryos. At E6 and E8,
cFgfr4 was found within the basilar papilla, but many more transcripts were
found surrounding the cochlear duct.
Overall, the role of Fgf19 in neural side fate of the basilar papilla
was not determined. Possible reasons for
the lack of phenotypic changes include nonfunctional Fgf19 being secreted which
could not bind and induce downstream signaling, Fgf19 being responsible for an
untested aspect of the cWnt9a overexpression model, or other misregulated genes
would be needed for the phenotypic change to occur.</p>
| Identifer | oai:union.ndltd.org:purdue.edu/oai:figshare.com:article/13643792 |
| Date | 01 March 2021 |
| Creators | Elizabeth Wehren (10035161) |
| Source Sets | Purdue University |
| Detected Language | English |
| Type | Text, Thesis |
| Rights | CC BY 4.0 |
| Relation | https://figshare.com/articles/thesis/EXPLORING_THE_ROLE_OF_FGFS_ON_RADIAL_PATTERNING_OF_THE_EMBRYONIC_CHICKEN_COCHLEA/13643792 |
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