The developing brain is sensitive to chemical exposures, however, currently there is a lack of good, regulatory accepted methods to study developmental neurotoxicity (DNT). The cell line C17.2 can be used as a model for DNT studies since it has the capacity to differentiate into neuronal and neuroglial populations. However, in a recent experiment, single cell RNA sequencing (scRNAseq) indicated that the cell line might contain a good proportion of fibroblasts. The aim of this project was to complement the scRNAseq with staining for protein markers for specific cell types in order to elucidate the cellular composition of differentiated and undifferentiated C17.2 cultures. The markers Hyaluronan-Mediated Motility Receptor (HMMR), vimentin and Platelet Derived Growth Factor Receptor A (PDGFRA) were identified as promising markers for radial glial cells (RGC) and subsequently fibroblast and validated by immunofluorescence. HMMR by itself was used as a marker for RGC and was, as expected, decreasing during the differentiation process. Co-localization of vimentin and PDGFRA was used as markers for fibroblast. However, the conditions for the PDGFRA-antibody could not be optimized due to time restraints. Thus no specific staining could be obtained and no conclusions could be drawn regarding the presence or absence of fibroblasts in the culture. The results emphasize the need for more optimisation or the selection of more specific markers, e.g. Collagen type I alpha 1 (Col1a1). Further, these findings highlight the complexity of the cellular composition and the need for other methods to characterize C17.2.
| Identifer | oai:union.ndltd.org:UPSALLA1/oai:DiVA.org:uu-521706 |
| Date | January 2024 |
| Creators | Thunegard, Lisa |
| Publisher | Uppsala universitet, Fysiologi och miljötoxikologi, Uppsala universitet, Institutionen för biologisk grundutbildning |
| Source Sets | DiVA Archive at Upsalla University |
| Language | English |
| Detected Language | English |
| Type | Student thesis, info:eu-repo/semantics/bachelorThesis, text |
| Format | application/pdf |
| Rights | info:eu-repo/semantics/openAccess |
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