The primary cilium is an important signaling organelle present on the membrane of almost all cell types during the G0 phase of the cell cycle. In addition to it signaling function, bending of primary cilia allows the cell to sense surrounding fluid flow, important for proper kidney function, as well as left and right direction-- features required during organ development. Cilia are involved in critical signaling functions found within the Hedgehog and PDGFRα pathways. The absence of cilia on the cell surface can significantly alter disease progression and drug response. Many common human ailments, including inflammation, tumorigenesis, and tissue aging, are often associated with changes in the extracellular matrix (ECM) properties, including an increase in ECM stiffness. We wondered whether ECM properties can also affect ciliation. While few past studies have noted that ciliation is affected by a change in substrate stiffness, a deeper insight on the role of mechanical forces from the extracellular matrix (ECM) in ciliogenesis is missing. This study aims to start unveiling the link between mechanical properties of ECM and ciliogenesis, with a focus on cilia signaling and extracellular cues responsible for cilia assembly and disassembly. To address how mechanics of ECM affects cilia biogenesis, we prepared gelatin substrates with various cross-linking degrees and measured ciliation level, loss of cilia, and cilia length. To visualize and measure the cilia, we used immunofluorescent labeling followed by fluorescent and confocal microscopy of the samples.
This study displays that increasing the degree of ECM cross-linking promotes ciliogenesis under serum deprivation. Higher cross-linking degree of gelatin substrates resulted in a higher ciliation rate as well as cilia elongation within the cell population. When cilia disassembly is initiated by the addition of serum-enriched medium, cells grown on highly cross-linked gelatin show higher ciliation rate when compared to cells grown on non-cross-linked gelatin. Expression level of cilia-associated protein acetylated α-tubulin was also higher on the highly cross-linked gelatin substrate. The final part of the study describes the action of cilia-targeting drugs in context of ECM cross-linking degree. The performance of ganetespib, a common cancer therapeutic and HSP90 inhibitor, shows a decrease in efficacy on substrate with high degree of cross-linking. / Biomedical Sciences
| Identifer | oai:union.ndltd.org:TEMPLE/oai:scholarshare.temple.edu:20.500.12613/6475 |
| Date | January 2021 |
| Creators | Belova, Elizaveta |
| Contributors | Golemis, Erica, Gligorijevic, Bojana, Tuzel, Erkan |
| Publisher | Temple University. Libraries |
| Source Sets | Temple University |
| Language | English |
| Detected Language | English |
| Type | Thesis/Dissertation, Text |
| Format | 40 pages |
| Rights | IN COPYRIGHT- This Rights Statement can be used for an Item that is in copyright. Using this statement implies that the organization making this Item available has determined that the Item is in copyright and either is the rights-holder, has obtained permission from the rights-holder(s) to make their Work(s) available, or makes the Item available under an exception or limitation to copyright (including Fair Use) that entitles it to make the Item available., http://rightsstatements.org/vocab/InC/1.0/ |
| Relation | http://dx.doi.org/10.34944/dspace/6457, Theses and Dissertations |
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