Towards Reduction or Substitution of Cytotoxic DMSO in Biobanking of Functional Bioengineered Megakaryocytes
Donor platelet transfusion is currently the only efficient treatment of life-threatening thrombocytopenia, but it is highly challenged by immunological, quality, and contamination issues, as well as short shelf life of the donor material. Ex vivo produced megakaryocytes and platelets represent a pro...
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Published in | International journal of molecular sciences Vol. 21; no. 20; p. 7654 |
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Main Authors | , , , , , , , |
Format | Journal Article |
Language | English |
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16.10.2020
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Abstract | Donor platelet transfusion is currently the only efficient treatment of life-threatening thrombocytopenia, but it is highly challenged by immunological, quality, and contamination issues, as well as short shelf life of the donor material. Ex vivo produced megakaryocytes and platelets represent a promising alternative strategy to the conventional platelet transfusion. However, practical implementation of such strategy demands availability of reliable biobanking techniques, which would permit eliminating continuous cell culture maintenance, ensure time for quality testing, enable stock management and logistics, as well as availability in a ready-to-use manner. At the same time, protocols applying DMSO-based cryopreservation media were associated with increased risks of adverse long-term side effects after patient use. Here, we show the possibility to develop cryopreservation techniques for iPSC-derived megakaryocytes under defined xeno-free conditions with significant reduction or complete elimination of DMSO. Comprehensive phenotypic and functional in vitro characterization of megakaryocytes has been performed before and after cryopreservation. Megakaryocytes cryopreserved DMSO-free, or using low DMSO concentrations, showed the capability to produce platelets in vivo after transfusion in a mouse model. These findings propose biobanking approaches essential for development of megakaryocyte-based replacement and regenerative therapies. |
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AbstractList | Donor platelet transfusion is currently the only efficient treatment of life-threatening thrombocytopenia, but it is highly challenged by immunological, quality, and contamination issues, as well as short shelf life of the donor material. Ex vivo produced megakaryocytes and platelets represent a promising alternative strategy to the conventional platelet transfusion. However, practical implementation of such strategy demands availability of reliable biobanking techniques, which would permit eliminating continuous cell culture maintenance, ensure time for quality testing, enable stock management and logistics, as well as availability in a ready-to-use manner. At the same time, protocols applying DMSO-based cryopreservation media were associated with increased risks of adverse long-term side effects after patient use. Here, we show the possibility to develop cryopreservation techniques for iPSC-derived megakaryocytes under defined xeno-free conditions with significant reduction or complete elimination of DMSO. Comprehensive phenotypic and functional in vitro characterization of megakaryocytes has been performed before and after cryopreservation. Megakaryocytes cryopreserved DMSO-free, or using low DMSO concentrations, showed the capability to produce platelets in vivo after transfusion in a mouse model. These findings propose biobanking approaches essential for development of megakaryocyte-based replacement and regenerative therapies. |
Author | Gryshkov, Oleksandr Eicke, Dorothee Wolkers, Willem F Figueiredo, Constança Schulze, Kai Pogozhykh, Denys Guzmán, Carlos A Blasczyk, Rainer |
AuthorAffiliation | 3 Unit for Reproductive Medicine, University of Veterinary Medicine Hannover, 30559 Hannover, Germany; Willem.Frederik.Wolkers@tiho-hannover.de 1 Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany; Dorothee.Eicke@gmx.de (D.E.); Blasczyk.Rainer@mh-hannover.de (R.B.) 4 Department of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany; Kai.Schulze@helmholtz-hzi.de (K.S.); CarlosAlberto.Guzman@helmholtz-hzi.de (C.A.G.) 2 Institute for Multiphase Processes, Leibniz Universität Hannover, 30823 Garbsen, Germany; Gryshkov@imp.uni-hannover.de |
AuthorAffiliation_xml | – name: 2 Institute for Multiphase Processes, Leibniz Universität Hannover, 30823 Garbsen, Germany; Gryshkov@imp.uni-hannover.de – name: 3 Unit for Reproductive Medicine, University of Veterinary Medicine Hannover, 30559 Hannover, Germany; Willem.Frederik.Wolkers@tiho-hannover.de – name: 4 Department of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany; Kai.Schulze@helmholtz-hzi.de (K.S.); CarlosAlberto.Guzman@helmholtz-hzi.de (C.A.G.) – name: 1 Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany; Dorothee.Eicke@gmx.de (D.E.); Blasczyk.Rainer@mh-hannover.de (R.B.) |
Author_xml | – sequence: 1 givenname: Denys orcidid: 0000-0003-1086-5190 surname: Pogozhykh fullname: Pogozhykh, Denys organization: Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany – sequence: 2 givenname: Dorothee surname: Eicke fullname: Eicke, Dorothee organization: Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany – sequence: 3 givenname: Oleksandr surname: Gryshkov fullname: Gryshkov, Oleksandr organization: Institute for Multiphase Processes, Leibniz Universität Hannover, 30823 Garbsen, Germany – sequence: 4 givenname: Willem F orcidid: 0000-0002-7406-4651 surname: Wolkers fullname: Wolkers, Willem F organization: Unit for Reproductive Medicine, University of Veterinary Medicine Hannover, 30559 Hannover, Germany – sequence: 5 givenname: Kai orcidid: 0000-0003-2286-3416 surname: Schulze fullname: Schulze, Kai organization: Department of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany – sequence: 6 givenname: Carlos A orcidid: 0000-0002-2913-5254 surname: Guzmán fullname: Guzmán, Carlos A organization: Department of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany – sequence: 7 givenname: Rainer orcidid: 0000-0003-3875-3190 surname: Blasczyk fullname: Blasczyk, Rainer organization: Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany – sequence: 8 givenname: Constança orcidid: 0000-0003-2786-0388 surname: Figueiredo fullname: Figueiredo, Constança organization: Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, 30625 Hannover, Germany |
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Keywords | propane-1,2-diol dimethyl sulfoxide megakaryocytes biobanking mouse model ethylene glycol induced pluripotent stem cells (iPSC) transfusion cytotoxicity platelets |
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SubjectTerms | Animals Apoptosis Availability biobanking Biobanks Bioengineering Blood Platelets - cytology Blood Platelets - drug effects Blood Preservation - methods Bone marrow Cell culture Cells, Cultured Contamination Cryopreservation Cryoprotective Agents - toxicity Cytotoxicity Dimethyl Sulfoxide - toxicity Donor materials Good Manufacturing Practice Humans Immunology In vivo methods and tests induced pluripotent stem cells (iPSC) Induced Pluripotent Stem Cells - cytology Induced Pluripotent Stem Cells - drug effects Logistics Megakaryocytes Megakaryocytes - cytology Megakaryocytes - drug effects Mice Mice, SCID mouse model platelets Population Shelf life Thrombocytopenia Transfusion |
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Title | Towards Reduction or Substitution of Cytotoxic DMSO in Biobanking of Functional Bioengineered Megakaryocytes |
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