@phdthesis{Dinger2008,
  author    = {Dinger, Timo Christoph},
  title     = {Neural Differentiation Potential of Murine Androgenetic Embryonic Stem Cells},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-36215},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2008},
  abstract  = {Uniparental zygotes with two genomes from the same sex can be established from fertilised oocytes after pronuclear exchange. They contain two maternal (gynogenetic; GG) or paternal (androgenetic; AG) pronuclei and are not competent to develop into viable offspring but they can form blastocysts from which embryonic stem cells (ES cells) can be derived. The developmental potential of uniparental ES cells is not fully investigated. The restricted developmental potential of uniparental cells is cell-intrinsic and probably reflects the different roles maternal and paternal genomes play during development. Following blastocyst injection, both GG and AG ES cells show biased and parent-of-origin-specific chimaera formation. While the in vitro and in vivo neural differentiation potential of GG ES cells is well characterised the neural developmental potential of AG ES cells is less clear. In an earlier study the group of K. John McLaughlin reported that AG and GG ES cell-derived hematopoietic stem cells conveyed long-term, multi-lineage hematopoietic engraftment with no associated pathologies (Eckardt et al., 2007). The aim of this study was to investigate the potential of AG uniparental murine ES cells to differentiate in vitro and in vivo into neural progenitor / stem cells and further into neurons, astro- and oligodendroglia in comparison to GG and biparental (normal fertilised; N) ES cells. Uniparental and biparental ES cells were obtained from K. John McLaughlin's group and a cell culture system was established to expand uniparental (AG, GG) and biparental N ES cells on murine embryonic fibroblasts (MEF). A multistep-protocol was used to differentiate ES cells towards pan-neural progenitor cells and neuronal and glial cell types (Br{\"u}stle et al., 1997). The ability of terminal neural differentiation in vitro was analysed by fluorescence microscopy using neuronal and glial lineage markers. In parallel, eGFP+ AG or N ES cells were injected into blastocysts prior to their transfer into foster mothers. At E12.5 and E14.5, embryos were isolated, forebrains were dissected and by means of fluorescence activated cell sorting (FACS) eGFP+ donor cells were isolated from chimeric brains. Both eGFP+ donor and corresponding eGFP- blastocyst-derived brain cells were expanded and analyses of differentiation potential and self-renewal capacity were performed. Also, cryosections of E12.5 chimeric brains were analysed for donor contribution to the neuronal lineage by immunofluorescence microscopy. Here it is described that following in vitro differentiation, AG pan-neural progenitor cells have similar abilities to differentiate into neuronal and glial lineages as GG and N pan-neural progenitor cells. In cryosections of E12.5 chimeric brains no differences in brain engraftment and formation of immature neuronal cells between uniparental AG and N donor cells were detected. AG and N ES cell-derived cells isolated from chimeric foetal brains by FACS exhibited similar neurosphere initiating cell frequencies and neural multi-lineage differentiation potential. Therefore, the data of this study suggest that the previously described differences in the in vivo engraftment pattern of uniparental inner cell mass (ICM) cells in foetal brains (Keverne et al., 1996) are not primarily due to limitations in the proliferation or differentiation properties of uniparental neural progenitor cells. The results presented here indicate that AG ES cell-derived neural progenitor / stem cells did not differ from N neural progenitor / stem cells in their self-renewal and their neural multi-lineage differentiation potential. Also AG ES cell-derived cells contributed to developing brains at early foetal developmental stages showing a widespread and balanced distribution in chimeric brains. AG brain cells form neurospheres with self-renewal and neural differentiation capacity similar to N ES cell-derived brain cells. Thus, the data of this study together indicate that the neural developmental potential in vivo and in vitro of AG and N ES cells does not differ.},
  subject      = {Stammzelle},
  language  = {en}
}
@phdthesis{Mueller2009,
  author    = {M{\"u}ller, Sophia},
  title     = {Molekulare Untersuchungen zur Rolle von Homeobox-Genen bei der Entwicklung von Echinococcus multilocularis},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-35616},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2009},
  abstract  = {Die alveol{\"a}re Echinokokkose ist eine, vorrangig in der n{\"o}rdlichen Hemisph{\"a}re verbreitete, parasit{\"a}re Erkrankung. Verursacht wird sie beim Menschen durch das Larvenstadium des Fuchsbandwurms. Homeoboxgene sind hochkonservierte Gene, die die Morphogenese von Lebewesen steuern. Die Anzahl und die Bedeutung von Homeoboxgenen in der Entwicklung von E.multilocularis waren bislang unbekannt. Im Rahmen dieser Arbeit konnten mit Hilfe von Sequenzanalysen im Genom des Fuchsbandwurms erstmals Homeoboxgene identifiziert und deren Expressionsmuster mittels PCR in verschiedenen Larvenstadien charakterisiert werden. Von insgesamt 23 gefundenen Homeoboxgenen wurden 15 Gene auf ihre larvenstadienspezifische Expression untersucht. Neun der untersuchten Gene zeigten in dem gew{\"a}hlten Versuchsaufbau eine Expression in den untersuchten Larvenstadien, f{\"u}nf davon zeigten eine verst{\"a}rkte Expression in den sp{\"a}ten Larvenstadien. F{\"u}r acht dieser neun Gene ließen sich dar{\"u}ber hinaus Hinweise auf eine Prozessierung ihrer mRNA {\"u}ber den Mechanismus des Trans-Spleißens finden. Vorangehende Versuche der Arbeitsgruppe von Prof. Brehm hatten einen Zusammenhang zwischen einer Stimulation fr{\"u}her Entwicklungsstadien der Parasitenlarven mit dem Zytokin BMP-2 und dessen rascherer Entwicklung in sp{\"a}tere Entwicklungsstadien nahegelegt. Die Auswirkung einer Behandlung fr{\"u}her Entwicklungsstadien mit dem Zytokin BMP-2 auf die jeweilige Genexpression wurde daher f{\"u}r die ausgew{\"a}hlten 15 Gene {\"u}berpr{\"u}ft. F{\"u}nf Gene zeigten unter dessen Einfluss eine verst{\"a}rkte Expression. Zwei darunter waren solche, die eine st{\"a}rkere Expression in sp{\"a}ten Larvenstadien aufwiesen. Diese zwei Gene stellen nun Kandidaten dar, die an der Entwicklung von E.multilocularis maßgeblich beteiligt sein k{\"o}nnten. Durch die Untersuchungen dieser Arbeit ergaben sich wichtige Hinweise auf die Entwicklung und die Regulationsmechanismen der Genexpression von E. multilocularis. Sie bilden eine Grundlage, die Rolle der Homeoboxgene f{\"u}r den Fuchsbandwurm n{\"a}her zu beschreiben und die hormonelle Kreuzregulation zwischen Parasit und Wirt weiter zu studieren.},
  subject      = {Fuchsbandwurm},
  language  = {de}
}
@phdthesis{Braasch2009,
  author    = {Braasch, Ingo},
  title     = {Evolution by genome duplication: insights from vertebrate neural crest signaling and pigmentation pathways in teleost fishes},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-35702},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2009},
  abstract  = {Gene and genome duplications are major mechanisms of eukaryotic genome evolution. Three rounds of genome duplication have occurred in the vertebrate lineage, two rounds (1R, 2R) during early vertebrate evolution and a third round, the fish-specific genome duplication (FSGD), in ray-finned fishes at the base of the teleost lineage. Whole genome duplications (WGDs) are considered to facilitate speciation processes and to provide the genetic raw material for major evolutionary transitions and increases in morphological complexity. In the present study, I have used comparative genomic approaches combining molecular phylogenetic reconstructions, synteny analyses as well as gene function studies (expression analyses and knockdown experiments) to investigate the evolutionary consequences and significance of the three vertebrate WGDs. First, the evolutionary history of the endothelin signaling system consisting of endothelin ligands and receptors was reconstructed. The endothelin system is a key component for the development of a major vertebrate innovation, the neural crest. This analysis shows that the endothelin system emerged in an ancestor of the vertebrate lineage and that its members in extant vertebrate genomes are derived from the vertebrate WGDs. Each round of WGD was followed by co-evolution of the expanding endothelin ligand and receptor repertoires. This supports the importance of genome duplications for the origin and diversification of the neural crest, but also underlines a major role for the co-option of new genes into the neural crest regulatory network. Next, I have studied the impact of the FSGD on the evolution of teleost pigment cell development and differentiation. The investigation of 128 genes showed that pigmentation genes have been preferentially retained in duplicate after the FSGD so that extant teleost genomes contain around 30\% more putative pigmentation genes than tetrapods. Large parts of pigment cell regulatory pathways are present in duplicate being potentially involved in teleost pigmentary innovations. There are also important differences in the retention of duplicated pigmentation genes among divergent teleost lineages. Functional studies of pigment synthesis enzymes in zebrafish and medaka, particularly of the tyrosinase family, revealed lineage-specific functional evolution of duplicated pigmentation genes in teleosts, but also pointed to anciently conserved gene functions in vertebrates. These results suggest that the FSGD has facilitated the evolution of the teleost pigmentary system, which is the most complex and diverse among vertebrates. In conclusion, the present study supports a major role of WGDs for phenotypic evolution and biodiversity in vertebrates, particularly in fish.},
  subject      = {Molekulare Evolution},
  language  = {en}
}
@phdthesis{Liedtke2007,
  author    = {Liedtke, Daniel},
  title     = {Functional divergence of Midkine growth factors : Non-redundant roles during neural crest induction, brain patterning and somitogenesis},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-25707},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2007},
  abstract  = {Neural crest cells and sensory neurons are two prominent cell populations which are induced at the border between neural and non-neural ectoderm during early vertebrate development. The neural crest cells are multipotent and highly migratory precursors that give rise to face cartilage, peripheral neurons, glia cells, pigment cells and many other cell types unique to vertebrates. Sensory neurons are located dorsally in the neural tube and are essential for sensing and converting environmental stimuli into electrical motor reflexes. In my PhD thesis, I obtained novel insights into the complex processes of cell induction at the neural plate border by investigating the regulation and function of mdkb in zebrafish. First, it was possible to demonstrate that mdkb expression is spatiotemporally correlated with the induction of neural crest cells and primary sensory neurons at the neural plate border. Second, it became evident that the expression of mdkb is activated by known neural crest cell inducing signals, like Wnts, FGFs and RA, but that it is independent of Delta-Notch signals essential for lateral inhibition. Knockdown experiments showed that mdkb function is necessary for induction of neural crest cells and sensory neurons at the neural plate border, probably through determination of a common pool of progenitor cells during gastrulation. The present study also used the advantages of the zebrafish model system to investigate the in vivo function of all midkine gene family members during early brain development. In contrast to the situation in mouse, all three zebrafish genes show distinct expression patterns throughout CNS development. mdka, mdkb and ptn expression is detected in mostly non-overlapping patterns during embryonic brain development in the telencephalon, the mid-hindbrain boundary and the rhombencephalon. The possibility of simultaneously knocking down two or even three mRNAs by injection of morpholino mixtures allowed the investigation of functional redundancy of midkine factors during brain formation. Knockdown of Midkine proteins revealed characteristic defects in brain patterning indicating their association with the establishment of prominent signaling centers such as the mid-hindbrain boundary and rhombomere 4. Interestingly, combined knockdown of mdka, mdkb and ptn or single knockdown of ptn alone prevented correct formation of somites, either by interfering with the shifting of the somite maturation front or interferance with cell adhesion in the PSM. Thus, Ptn was identified as a novel secreted regulator of segmentation in zebrafish.},
  subject      = {Zebrab{\"a}rbling},
  language  = {en}
}
@phdthesis{Kluever2007,
  author    = {Kl{\"u}ver, Nils},
  title     = {Molecular analysis of gonad development in medaka (Oryzias latipes) and Oryzias celebensis},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-25105},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2007},
  abstract  = {The process of sex-determination can be better understood through examinations of developing organs and cells, which are involved in the formation of undifferentiated gonad. This mechanisms show in fish a broad variety, ranging from hermaphroditism to gonochorism and environmental to genetic sex determination. Hormones and abiotic factors such as temperature and pH can influence teleost development and reproductive traits. These factors are vulnerable to pollutants and climate changes. Therefore, it is important to examine gonad development and sex-determination/differentiation in teleost fish. Teleost fish are the largest known group of vertebrates with approximately 25,000 species and are used for such kind of examinations as model organisms. Recently, in Oryzias latipes (medaka), dmrt1bY (or dmy), a member of the Dmrt gene family, has been described as testis-determining gene. However, this gene is not the universal master sex-determining gene in teleost fish. Although dmrt1bY is present in the most closely related species of the genus, namely Oryzias curvinotous, it is absent from other Oryzias species, like Oryzias celebensis, and other fish. During my thesis, I studied gonad development in medaka and in the closely related species Oryzias celebensis. Germ cell specification in medaka seems to be dependent on maternally provided cytoplasmatic determinants, so called germ plasm. Nanos and vasa are such germ cell specific genes. In zebrafish they are asymmetrically localized in the early embryo. I have shown that nanos mRNA is evenly distributed in the early embryo of medaka. A similar pattern has been already described for the medaka vasa homolog, olvas. This suggests differences in PGC specification in zebrafish and medaka. Further, the vasa homolog was isolated and the expression pattern examined in O. celebensis. The results show that it can be used as a germ cell specific marker. Additionally, the primordial germ cell migration in O. celebensis was followed, which is similar to medaka PGC migration. Primordial germ cell migration in vertebrates is dependent on the chemokine stromal cell-derived factor 1 (Sdf-1). Medaka has two different sdf-1 genes, sdf-1a and sdf-1b. Both genes are expressed in the lateral plate mesoderm (LPM). During late embryonic development, I could show that sdf-1a is expressed in newly formed somites and not longer in the LPM. Sdf-1b expression persisted in the posterior part of the lateral plate mesoderm in the developing gonad. In terms of early and late functions, this suggests subfunctionalization of sdf-1a and sdf-1b. In "higher" vertebrates, genes that are involved in the process of gonad development have been studied in detail, e.g. Wt1, Sox9, and Amh. I have analyzed the expression pattern of wt1 and sox9 co-orthologs and amh. In both, the medaka and O. celebensis, wt1a transcripts were localized in the LPM and its expression was similar to sdf-1a gene expression in medaka. Wt1b expression was restricted to the developing pronephric region. During later embryonic development, wt1a is specifically expressed in the somatic cells of the gonad primordium in both sexes. This is the first time that in fish wt1 gene expression in developing gonads has been described. Therefore, this result suggests that wt1a is involved in the formation of the bipotential gonad. Furthermore, I have analyzed the gonad specific function of the wt1 co-orthologs in medaka. I could show that a conditional co-regulation mechanism between Wt1a and Wt1b ensures PGC maintenance and/or survival. The expression of sox9 genes in medaka and sox9b in O. celebensis were detected in the somatic cells of the gonad primordium of both sexes. Additionally, I have shown that amh and amhrII in medaka are expressed in somatic cells of the gonad primordium of both sexes. This suggests that sox9b, amh and amhrII are involved in gonad development and have specific functions in the adult gonad. In O. celebensis I could detect an expression of dmrt1 already six days after fertilization in half of the embryos, which is similar to the dmrt1bY expression in medaka. Whether the expression of dmrt1 is male specific in O. celebensis is currently under investigation. Altogether, the obtained results provide new insights into gene expression patterns during the processes of gonad development. Furthermore, no differences in the expression pattern of wt1a and sox9b during gonad development between the medaka and O. celebensis could be detected. This might indicate that the genetic mechanisms during gonad development are similar in both species.},
  subject      = {Japank{\"a}rpfling},
  language  = {en}
}