@article{CullmannJahnSpindleretal.2021,
  author    = {Cullmann, Katharina and Jahn, Magdalena and Spindler, Markus and Schenk, Franziska and Manukjan, Georgi and Mucci, Adele and Steinemann, Doris and Boller, Klaus and Schulze, Harald and Bender, Markus and Moritz, Thomas and Modlich, Ute},
  title     = {Forming megakaryocytes from murine-induced pluripotent stem cells by the inducible overexpression of supporting factors},
  series = {Research and Practice in Thrombosis and Haemostasis},
  volume    = {5},
  journal   = {Research and Practice in Thrombosis and Haemostasis},
  number    = {1},
  doi       = {10.1002/rth2.12453},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-224565},
  pages     = {111 -- 124},
  year      = {2021},
  abstract  = {Background Platelets are small anucleate cells that circulate in the blood in a resting state but can be activated by external cues. In case of need, platelets from blood donors can be transfused. As an alternative source, platelets can be produced from induced pluripotent stem cells (iPSCs); however, recovered numbers are low. Objectives To optimize megakaryocyte (MK) and platelet output from murine iPSCs, we investigated overexpression of the transcription factors GATA-binding factor 1 (GATA1); nuclear factor, erythroid 2; and pre-B-cell leukemia transcription factor 1 (Pbx1) and a hyperactive variant of the small guanosine triphosphatase RhoA (RhoAhc). Methods To avoid off-target effects, we generated iPSCs carrying the reverse tetracycline-responsive transactivator M2 (rtTA-M2) in the Rosa26 locus and expressed the factors from Tet-inducible gammaretroviral vectors. Differentiation of iPSCs was initiated by embryoid body (EB) formation. After EB dissociation, early hematopoietic progenitors were enriched and cocultivated on OP9 feeder cells with thrombopoietin and stem cell factor to induce megakaryocyte (MK) differentiation. Results Overexpression of GATA1 and Pbx1 increased MK output 2- to 2.5-fold and allowed prolonged collection of MK. Cytologic and ultrastructural analyses identified typical MK with enlarged cells, multilobulated nuclei, granule structures, and an internal membrane system. However, GATA1 and Pbx1 expression did not improve MK maturation or platelet release, although in vitro-generated platelets were functional in spreading on fibrinogen or collagen-related peptide. Conclusion We demonstrate that the use of rtTA-M2 transgenic iPSCs transduced with Tet-inducible retroviral vectors allowed for gene expression at later time points during differentiation. With this strategy we could identify factors that increased in vitro MK production.},
  language  = {en}
}
@article{NguyenKraftYuetal.2015,
  author    = {Nguyen, Minh Thu and Kraft, Beatrice and Yu, Wenqi and Demicrioglu, Dogan Doruk and Hertlein, Tobias and Burian, Marc and Schmaler, Mathias and Boller, Klaus and Bekeredjian-Ding, Isabelle and Ohlsen, Knut and Schittek, Birgit and G{\"o}tz, Friedrich},
  title     = {The vSa\(\alpha\) Specific Lipoprotein Like Cluster (lpl) of S. aureus USA300 Contributes to Immune Stimulation and Invasion in Human Cells},
  series = {PLoS Pathogens},
  volume    = {11},
  journal   = {PLoS Pathogens},
  number    = {6},
  doi       = {10.1371/journal.ppat.1004984},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-151856},
  pages     = {e1004984},
  year      = {2015},
  abstract  = {All Staphylococcus aureus genomes contain a genomic island, which is termed vSa\(\alpha\) and characterized by two clusters of tandem repeat sequences, i.e. the exotoxin (set) and 'lipoprotein-like' genes (lpl). Based on their structural similarities the vSa\(\alpha\) islands have been classified as type I to IV. The genomes of highly pathogenic and particularly epidemic S. aureus strains (USA300, N315, Mu50, NCTC8325, Newman, COL, JH1 or JH9) belonging to the clonal complexes CC5 and CC8 bear a type I vSa\(\alpha\) island. Since the contribution of the lpl gene cluster encoded in the vSa\(\alpha\) island to virulence is unclear to date, we deleted the entire lpl gene cluster in S. aureus USA300. The results showed that the mutant was deficient in the stimulation of pro-inflammatory cytokines in human monocytes, macrophages and keratinocytes. Purified lipoprotein Lpl1 was further shown to elicit a TLR2-dependent response. Furthermore, heterologous expression of the USA300 lpl cluster in other S. aureus strains enhanced their immune stimulatory activity. Most importantly, the lpl cluster contributed to invasion of S. aureus into human keratinocytes and mouse skin and the non-invasive S. carnosus expressing the lpl gene cluster became invasive. Additionally, in a murine kidney abscess model the bacterial burden in the kidneys was higher in wild type than in mutant mice. In this infection model the lpl cluster, thus, contributes to virulence. The present report is one of the first studies addressing the role of the vSa\(\alpha\) encoded lpl gene cluster in staphylococcal virulence. The finding that the lpl gene cluster contributes to internalization into non-professional antigen presenting cells such as keratinocytes high-lights the lpl as a new cell surface component that triggers host cell invasion by S. aureus. Increased invasion in murine skin and an increased bacterial burden in a murine kidney abscess model suggest that the lpl gene cluster serves as an important virulence factor.},
  language  = {en}
}