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Browsing by Subject "developmental biology"

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  • Tervi, Anniina (2020)
    The diversity of different neuronal types lays the foundation for different functions in the brain. The development of different subpopulations and special features of neurons in the central nervous system are still partly unknown. Finding answers to these developmental issues could help in the process of characterisation of cell types and mapping of neuronal networks between the brainstem nuclei in the brain. Previous studies have shown that a ventrolateral neuroepithelial domain in the anterior hindbrain, rV2, produces excitatory (glutamatergic) and inhibitory (GABAergic) neurons, which are related to monoaminergic nuclei in the brainstem (Lahti et al., 2016). In this master’s thesis project, the development of a subpopulation of neurons expressing Gsc2 transcription factor in the interpeduncular nucleus was studied. This project was based on single-cell RNA sequencing results conducted in E13.5 mice. Predicted by single-cell RNA sequencing results, Gsc2 expressing cells are GABAergic interneurons and originate from the rV2 domain of the rhombomere 1 region in the hindbrain. Co-expression pattern with another transcription factor Sall3 with Gsc2 during development was also addressed in the study. Furthermore, the role of Notch signalling in the binary cell fate decision between GABAergic and the glutamatergic fate of rV2 neurons was investigated. Validation of single-cell RNA sequencing results was performed using in situ hybridisation and immunohistochemistry methods with mice embryos at the age of E12.5 and E15.5. This study verified previously shown origin of Gsc2 expressing cells to the rhombomere 1 region and in addition, showed that Gsc2 expressing cells are GABAergic. Co-expression pattern of Gsc2 with Sall3 neither in the rV2 domain nor in the interpeduncular nucleus was seen in our results. In the rV2 domain, the depletion of Notch signalling decreased the expression of differentiating GABAergic neurons. This indicates that Notch has a role in GABAergic neurotransmitter identity during the development of brainstem neurons in mice. Based on our results, Gsc2 could be used as a lineage marker for GABAergic interneurons originating from the rhombomere 1 region and as a marker for a subpopulation of the interpeduncular nucleus. Furthermore, results from the role of Notch signalling could help in discovering the mechanisms related to the determination of neurotransmitter identity in rV2 neurons. Further investigations, in different developmental time points and with additional markers, are needed to verify these results.
  • Ollonen, Joni (2020)
    The skull represents the most highly diversified and evolutionarily adapted anatomical aspect of metazoans, and its development and evolution have been a major driving force in the expansion of vertebrates. The evolution of skull and lower jaw bones have led to the adaptive radiation of jawed vertebrates, and skull tissues have changed rapidly over time and were finely tuned to meet functional and ecological demands with tremendous precision. Because of the long-lasting interest in conventional animal models, there is no general genetic or developmental model of skull evolution and diversity in vertebrates. Squamate reptiles represent the best model to study those aspects because of their key basal phylogenetic position within amniotes (i.e., mammals, birds, reptiles) and their exceptionally high levels of morphological variation (including their kinetic skulls). In particular, their lower jaw bones display tremendous variation. In order to assess this variation and the ecological and developmental factors connected to it, several methods from different fields of biology have to be used. In this study, morphometric, embryology and developmental approaches are used to investigate the ecological and developmental factors associated with the diversification of lower jaw bones in snakes and lizards. The shape diversity of squamate lower jaw bones was approached in a systematic way, using geometric morphometrics. Embryological methods were used to compare the embryonic stage of available squamate model animals at oviposition and to assess the order of ossification of embryo with earliest developmental stage at oviposition (bearded dragon, Pogona vitticeps). In addition, expression of major conserved candidate genes at different stages of lower jaw development (pharyngeal arches, mesenchyme patterning, ossification) were assessed in this species. The results indicate that the lower jaw bones of snakes versus lizards but also of fossorial squamates versus other habitats are significantly different. Heterochrony was also detected at both early stages (pharyngeal arche development at oviposition) and at the onset of ossification in lizards and snakes. Coherent with that, alterations in the expression pattern of Dlx genes in pharyngeal arches were observed in bearded dragon in comparison to earlier studies with mice, while other conserved markers of skeletogenesis were rather conserved. This analysis of the genotype and phenotype map of the reptilian skull provides some new insights into the development, origin and divergence of vertebrate tissues. The results will establish a good basis for future studies involving comparative developmental biology of bearded dragon. Future studies will offer excellent new opportunities to link craniofacial morphology, genetics/genomics and development to both ecological adaptation and evolutionary biology.
  • Ryhänen, Emma (2023)
    Rare mutations in the primate specific ZNF808 gene are a novel cause of pancreatic agenesis, a congenital developmental disorder that leads to neonatal diabetes. ZNF808 loss-of-function has been shown to lead to aberrant activation of regulatory MER11 elements, followed by upregulation of genes in proximity to these elements and increased expression of hepatic lineage markers. These findings suggest ZNF808 to play a key role in balancing the differentiation of endoderm progenitor cells between pancreatic and liver lineages during early human development. This thesis work aimed to study the gene regulatory mechanisms of ZNF808 in the differentiating endoderm progenitor cells to understand its function in controlling pancreatic lineage specification. This was achieved by comparing the lineage specification processes in wild-type (H1) and ZNF808 knockout (H1-ZNF808-KO) human embryonic stem cells (hESCs) during pancreatic differentiation. Further characterization of cellular heterogeneity and gene expression profiles upon ZNF808 loss was done using single-cell RNA sequencing (scRNA-seq). To validate the role of ZNF808 as the mediator of the observed lineage specification bias, the phenotype rescue was examined in a ZNF808 knockout overexpression cell line (H1-ZNF808-KO-OX). The results of this study demonstrate a clear lineage specification bias in the ZNF808 knockout, seen as divergence of the multipotent endoderm progenitors towards alternate hepatic and biliary fates at the posterior foregut stage. By modifying the pancreatic differentiation protocol, we were able to observe phenotype manifestation and cellular heterogeneity suppressed in the standard differentiation conditions. The scRNA-seq data analysis revealed the emergence of a biliary cell population showing upregulation of several hepatic markers, suggesting an alternative lineage specification process governed by ZNF808. Additionally, preliminary results from ZNF808 overexpression showed rescue of the ZNF808 knockout phenotype, further supporting its critical role in the normal pancreatic lineage development. In conclusion, these findings demonstrate the important role of ZNF808 in early human pancreatic development and warrant further studies on the detailed gene regulatory network guiding pancreatic lineage specification.