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In-vitro-Charakterisierung und kardiale Differenzierung von induziert pluripotenten Stammzellen der Maus

dc.contributor.advisorBecker, Alexander Dr.
dc.contributor.authorLentzen, Max-Philipp
dc.titleIn-vitro-Charakterisierung und kardiale Differenzierung von induziert pluripotenten Stammzellen der Mausde
dc.title.translatedIn vitro characterisation and cardiac differentiation of murine induced pluripotent stem cellsde
dc.contributor.refereeGuan-Schmidt, Kaomei Prof. Dr.
dc.description.abstractengThe following paper is part of many recent research projects that try to find and establish a sufficient cell replacement therapy for structural myocardial deseases. Induced pluripotent stem cells which are comparable to embryonic stem cells and differentiate into cardiomyocytes seem to be an option. The task was to compare miPS with mES by characterisation and differentiation into cardiomyocytes and also characterise them. This was followed by antibiotic selection and purification via immunofluorescence. The in-vitro-results showed a successfull reprogramming with the transcription factors OCT4, SOX2, KLF4 und c-MYC in a single lentiviral stem cell cassette and also that the investigated miPS showed similar characteristics as the investigated ES. The proof of pluripotency showed no differences between the embryonic and the induced pluripotent stem cells neither on mRNA nor on protein level. The morphology of the cells during the differentiation of the cells of the three germ layers showed only minimal differences. There was also a physiological caryotype shown even after long term cultivation. By using a standardized EB based differentiation protocol (hanging drop-method) for ES the induced pluripotent and embryonic stem cells have been differentiated into cardiomyocytes over 22 days. The source of the investigated cells were double transgene mice (Mhc-Neo/Mhc-eGfp). With them it was possible to achieve a mhc dependent expression of eGfp and a neomycine resistance of the cardiomyocytes which made a selection and visualisation possible. Analysis of the molecular, structural and functional aspects of the iPS derived cardiomyocytes showed typical characteristics of ES derived cardiomyocytes. Investigations by rt-PCR showed the expression of typical mesodermal marker genes as Gata4, Nkx2.5, α-Mhc, Mlc2v and Anf. Also on protein level the expression of cardiac markers could be proofed. Thus the differentiation of induced pluripotent stem cells into functional cardiomyocytes could be demonstrated. By Use of iPS derived and not ES derived cardiomyocytes problems as immunological reactions or embryo consuming research as an ethical issue could be solved. Based on the results of this research project following investigations have to show if the generated cardiomyocytes are suitable for transplantation, if they survive and if they functional integrate in vivo. For a therapeutical application the achieved results also have to be reproduced in
dc.contributor.coRefereeDressel, Ralf Prof. Dr.
dc.subject.enginduced pluripotent stem cellsde
dc.subject.engembryonic stem cellsde
dc.subject.engtranscription factorsde
dc.subject.englentiviral stem cell cassettede
dc.subject.enghanging dropde
dc.subject.engdouble transgene micede
dc.affiliation.instituteMedizinische Fakultätde
dc.subject.gokfullMedizin (PPN619874732)de
dc.subject.gokfullInnere Medizin - Allgemein- und Gesamtdarstellungen (PPN619875747)de

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