@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{LauknerTruchetManukjanetal.2021,
  author    = {Laukner, Anna and Truchet, Laura and Manukjan, Georgi and Schulze, Harald and Langbein-Detsch, Ines and Mueller, Elisabeth and Leeb, Tosso and Kehl, Alexandra},
  title     = {Effects of cocoa genotypes on coat color, platelets and coagulation parameters in French Bulldogs},
  series = {Genes},
  volume    = {12},
  journal   = {Genes},
  number    = {7},
  issn      = {2073-4425},
  doi       = {10.3390/genes12071092},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-242745},
  year      = {2021},
  abstract  = {A nonsense variant in HPS3, c.2420G>A or p.Trp807*, was recently discovered as the cause for a brown coat color termed cocoa in French Bulldogs. Here, we studied the genotype-phenotype correlation regarding coat color in HPS3 mutant dogs that carried various combinations of mutant alleles at other coat color genes. Different combinations of HPS3, MLPH and TYRP1 genotypes resulted in subtly different shades of brown coat colors. As HPS3 variants in humans cause the Hermansky-Pudlak syndrome type 3, which in addition to oculocutaneous albinism is characterized by a storage pool deficiency leading to bleeding tendency, we also investigated the phenotypic consequences of the HPS3 variant in French Bulldogs on hematological parameters. HPS3 mutant dogs had a significantly lowered platelet dense granules abundance. However, no increased bleeding tendencies in daily routine were reported by dog owners. We therefore conclude that in dogs, the phenotypic effect of the HPS3 variant is largely restricted to pigmentation. While an effect on platelet morphology is evident, we did not obtain any indications for major health problems associated with the cocoa coat color in French Bulldogs. Further studies will be necessary to definitely rule out very subtle effects on visual acuity or a clinically relevant bleeding disorder.},
  language  = {en}
}