Browsing by Subject "developmental biology"

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  • Jia, S.; Zhou, J.; Wee, Y.; Mikkola, M. L.; Schneider, P.; D'Souza, R. N. (2017)
    To date, surgical interventions are the only means by which craniofacial anomalies can be corrected so that function, esthetics, and the sense of well-being are restored in affected individuals. Unfortunately, for patients with cleft palate-one of the most common of congenital birth defects-treatment following surgery is prolonged over a lifetime and often involves multidisciplinary regimens. Hence, there is a need to understand the molecular pathways that control palatogenesis and to translate such information for the development of noninvasive therapies that can either prevent or correct cleft palates in humans. Here, we use the well-characterized model of the Pax9(-/-) mouse, which displays a consistent phenotype of a secondary cleft palate, to test a novel therapeutic. Specifically, we demonstrate that the controlled intravenous delivery of a novel mouse monoclonal antibody replacement therapy, which acts as an agonist for the ectodysplasin (Eda) pathway, can resolve cleft palate defects in Pax9(-/-) embryos in utero. Such pharmacological interventions did not reverse the arrest in tooth, thymus, and parathyroid gland development, suggesting that the relationship of Pax9 to the Eda/Edar pathway is both unique and essential for palatogenesis. Expression analyses and unbiased gene expression profiling studies offer a molecular explanation for the resolution of palatal defects, showing that Eda and Edar-related genes are expressed in normal palatal tissues and that the Eda/Edar signaling pathway is downstream of Pax9 in palatogenesis. Taken together, our data uncover a unique relationship between Pax9 and the Eda/Edar signaling pathway that can be further exploited for the development of noninvasive, safe, and effective therapies for the treatment of cleft palate conditions and other single-gene disorders affecting the craniofacial complex.
  • Tervi, Anniina (Helsingin yliopisto, 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 (Helsingin yliopisto, 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.
  • Xu, Hongen; Pausch, Hubert; Venhoranta, Heli; Rutkowska, Karolina; Wurmser, Christine; Rieblinger, Beate; Flisikowska, Tatiana; Frishman, Dmitrij; Zwierzchowski, Lech; Fries, Ruedi; Andersson, Magnus; Kind, Alexander; Schnieke, Angelika; Flisikowski, Krzysztof (2017)
    Intrauterine growth restriction (IUGR) is caused by dysregulation of placental metabolism. Paternally inherited IUGR mutations in the fetus influence maternal physiology via the placenta. However, it is not known whether the maternal placenta also affects the extent of IUGR in such fetuses. In cattle and other ruminants, maternal-fetal communication occurs primarily at the placentomes. We previously identified a 3 deletion in the noncoding MER1 repeat containing imprinted transcript 1 (MIMT1) gene that, when inherited from the sire, causes IUGR and late abortion in Ayshire cattle with variable levels of severity. Here, we compared the transcriptome and genomic imprinting in fetal and maternal placentome components of wild-type and MIMT1(Del/WT) fetuses before IUGR became apparent, to identify key early events. Transcriptome analysis revealed fewer differentially expressed genes in maternal than fetal MIMT1(Del/WT) placentome. AST1, within the PEG3 domain, was the only gene consistently reduced in IUGR in both fetal and maternal samples. Several genes showed an imprinting pattern associated with IUGR, of which only secernin 3 (SCRN3) and paternally expressed 3 (PEG3) were differentially imprinted in both placentome components. Loss of strictly monoallelic, allele-specific expression (similar to 80:20) of PEG3 in the maternal MIMT1(Del/WT) placenta could be associated with incomplete penetrance of MIMT1(Del). Our data show that dysregulation of the PEG3 domain is involved in IUGR, but also reveal that maternal placental tissues may affect the penetrance of the paternally inherited IUGR mutation.
  • Veistinen, Lotta (Helsingin yliopisto, 2015)
    The flat bones of the skull, the calvarial bones, develop by intramembranous ossification during which mesenchymal cells first condense and subsequently differentiate into osteoblasts. Sutures separate the calvarial bones and facilitate the synchronized growth of the underlying brain and the calvaria. Hedgehog (Hh) signalling has an indisputable role in craniofacial development as well as during endochondral ossification. Yet, little is known about its function during intramembranous ossification of the calvarial bones. GLI-Kruppel family member 3 (Gli3) is a zinc-finger transcription factor that mediates Hh signalling. In the absence of Hh ligand Gli3 is proteolytically cleaved into a repressor that inhibits transcription of Hh target genes. Mutations in GLI3 cause Greig cephalopolysyndactyly syndrome in humans, in which an infrequent, but significant feature is premature fusion of the metopic suture (interfrontal suture in mice). We have used Gli3 loss-of- function mouse (Gli3Xt-J/Xt-J) as a model to investigate the effects of aberrant Hh signalling during calvarial development. In my thesis I describe how loss of Gli3 causes craniosynostosis of the lambdoid as well as interfrontal sutures in mice. Elevated proliferation and ectopic differentiation of osteoprogenitors underlies this phenomenon. We were able to rescue craniosynostosis in these mice by two mechanisms. Firstly, by elevating fibroblast growth factor (Fgf) signalling in the suture prior to its fusion by imbedding Fgf2 soaked beads in tissue culture. This induced Twist1 expression, which inhibits function of ectopically expressed Runx2. Secondly, craniosynostosis was prevented by genetically reducing Runx2 activity by generating Gli3Xt-J/Xt-J;Runx2+/- mice, which normalized elevated levels of Bmp signalling in the affected sutures. We also put forward a model of how Hh signalling helps to maintain the integrity of bone margins during calvarial development. The repressor isoform of Gli3 inhibits Runx2 activity in the early osteoprogenitor cells. Runx2, on the other hand, activates Ihh expression in the mature osteoblasts, which then induces osteogenesis by inhibiting the function of Gli3 repressor. Our findings indicate that Gli3 and Hh signalling have an important role in mediating the location of osteoblast differentiation and the speed of bone formation in the developing calvaria. Uncovering the cellular and molecular mechanisms that underlie normal calvarial development, as well as pathological processes, is a vital step in developing treatment strategies for patients with craniosynostosis.
  • Matsuda, Shinya (Helsingin yliopisto, 2013)
    Evolutionarily conserved signaling pathways mediate cell-cell communications during development. While the extracellular signal is precisely regulated to achieve dynamic morphogenetic events at the species level, it must be also flexible to generate the diversified morphologies through evolution. However, little is known about the mechanisms behind the precision and flexibility. The insect wing vein pattern can provide an excellent model to address this fundamental question, since species-specific wing vein patterns have been diversified through evolution. In this thesis, I study how evolutionarily conserved bone morphogenetic protein (BMP)-type ligand specifies diversified insect wing vein patterns using Dipteran Drosophila melanogaster and Hymenopteran sawfly Athalia rosae as models. In Drosophila, BMP-type ligand Decapentaplegic (Dpp) is expressed in the longitudinal veins (LVs) to maintain LVs and induce crossveins (CVs) fates during pupal stages. However, the distribution of Dpp remained largely unknown. Using GFP-tagged Dpp, I demonstrated that Dpp is directionally transported from LVs into the posterior crossvein (PCV) region by two extracellular BMP-binding proteins, Short gastrulation (Sog) and Crossveinless (Cv). In contrast, most of Dpp did not diffuse from LVs by Type I BMP receptor and a positive feedback mechanism. Thus the active transport and retention mechanisms allow diffusible Dpp to draw the complex wing vein patterns in Drosophila. To investigate how BMP signal instructs wing vein morphogenesis that involves apposition and cell shape changes between two wing epithelial layers, I then focused on the function of RhoGAP Crossveinless-C (Cv-C) during the PCV morphogenesis. I found that cv-c mediates PCV morphogenesis downstream of BMP signal by inactivating various Rho-type small GTPases. Interestingly, I found that cv-c is also required for Dpp transport, while Sog/Cv mediated BMP signal is guided at the ectopic wing veins caused by loss of Rho-type small GTPases. These observations identified a feed-forward mechanism coupling Dpp transport and PCV morphogenesis. To address how BMP signal specifies diversified insect wing vein patterns, I then introduced sawfly as a new model. I found that dpp is ubiquitously expressed but BMP signal reflects distinct fore- and hindwing vein patterns in sawfly. To address if Dpp transport mechanism is involved in wing vein formation, Cv/Tsg was identified from sawfly. Loss of dpp or cv/tsg by RNAi affected BMP signal and all of wing venations. These observations suggest that ubiquitously expressed Dpp is redistributed to specify distinct fore- and hindwing vein patterns in sawfly. Taken together, I found that the extracellular distribution of Dpp/BMP is tightly regulated and coordinated to achieve precise patterning and morphogenesis of the insect wing veins. Furthermore, this study raises an interesting possibility that changes in the directionality of Dpp/BMP diffusion may underlie distinct insect wing vein patterns.