Novel Method Creates Megakaryocytes From Stem Cells For Transfusion
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By LabMedica International staff writers Posted on 28 Apr 2016 |
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Image: The Axiovert 40 fluorescent microscope (Photo courtesy of Zeiss).
The production of megakaryocytes (MKs), the precursors of blood platelet, from human pluripotent stem cells (hPSCs) offers exciting clinical opportunities for transfusion medicine. Up to four components can be derived from donated blood: red cells, white cells, plasma and platelets.
Each component serves a different medical need, allowing several patients to benefit from a single unit of donation. Platelet transfusions are given to patients with life-threatening bleeding due to injury or surgery. They may also be given to patients having treatments for cancer or leukemia, or with blood disorders where they cannot make enough platelets of their own.
A large team of scientists led by those at the University of Cambridge and NHS Blood and Transplant (Cambridge, UK) developed an original approach for the large-scale generation of MKs in chemically defined conditions using a forward programming strategy relying on the concurrent exogenous expression of three transcription factors: globin transcription factor 1(GATA1,) Friend leukemia integration 1 transcription factor (FLI1) and T-Cell Acute Lymphocytic Leukemia 1 (TAL1).
A multiplicity of methodologies were used that included human pluripotent stem cell (hPSC) culture, selecting transcription factor candidates, transcription factor cloning using recombinant lentiviral vectors, human pluripotent stem cell transduction, megakaryocyte forward programming and flow cytometry analyses were performed on a CyAn ADP analyzer (Beckman Coulter, Brea, CA, USA). Megakaryocyte colony forming assay, and immunofluorescence analysis and the latter were visualized on an Axiovert 40 fluorescent microscope (Zeiss, Cambridge, UK).
The forward programmed MKs proliferated and differentiated in culture for several months with MK purity over 90% reaching up to 2 × 105 mature MKs per input hPSC. Functional platelets are generated throughout the culture allowing the prospective collection of several transfusion units from as few as one million starting hPSCs. The high cell purity and yield achieved by MK forward programming, combined with efficient cryopreservation and good manufacturing practice (GMP)-compatible culture, make this approach eminently suitable to both in vitro production of platelets for transfusion.
The authors noted that critically, the forward programmed MKs (fopMKs) matured into platelet-producing cells that could be cryopreserved, maintained and amplified in vitro for over 90 days showing an average yield of 200,000 MKs per input hPSC. The study was published on April 7, 2016, in the journal Nature Communications.
Related Links:
University of Cambridge and NHS Blood and Transplant
Beckman Coulter
Zeiss
Each component serves a different medical need, allowing several patients to benefit from a single unit of donation. Platelet transfusions are given to patients with life-threatening bleeding due to injury or surgery. They may also be given to patients having treatments for cancer or leukemia, or with blood disorders where they cannot make enough platelets of their own.
A large team of scientists led by those at the University of Cambridge and NHS Blood and Transplant (Cambridge, UK) developed an original approach for the large-scale generation of MKs in chemically defined conditions using a forward programming strategy relying on the concurrent exogenous expression of three transcription factors: globin transcription factor 1(GATA1,) Friend leukemia integration 1 transcription factor (FLI1) and T-Cell Acute Lymphocytic Leukemia 1 (TAL1).
A multiplicity of methodologies were used that included human pluripotent stem cell (hPSC) culture, selecting transcription factor candidates, transcription factor cloning using recombinant lentiviral vectors, human pluripotent stem cell transduction, megakaryocyte forward programming and flow cytometry analyses were performed on a CyAn ADP analyzer (Beckman Coulter, Brea, CA, USA). Megakaryocyte colony forming assay, and immunofluorescence analysis and the latter were visualized on an Axiovert 40 fluorescent microscope (Zeiss, Cambridge, UK).
The forward programmed MKs proliferated and differentiated in culture for several months with MK purity over 90% reaching up to 2 × 105 mature MKs per input hPSC. Functional platelets are generated throughout the culture allowing the prospective collection of several transfusion units from as few as one million starting hPSCs. The high cell purity and yield achieved by MK forward programming, combined with efficient cryopreservation and good manufacturing practice (GMP)-compatible culture, make this approach eminently suitable to both in vitro production of platelets for transfusion.
The authors noted that critically, the forward programmed MKs (fopMKs) matured into platelet-producing cells that could be cryopreserved, maintained and amplified in vitro for over 90 days showing an average yield of 200,000 MKs per input hPSC. The study was published on April 7, 2016, in the journal Nature Communications.
Related Links:
University of Cambridge and NHS Blood and Transplant
Beckman Coulter
Zeiss
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