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A Novel High-Throughput Screening Platform Reveals an Optimized Cytokine Formulation for Human Hematopoietic Progenitor Cell Expansion.

ABSTRACT: The main limitations of hematopoietic cord blood (CB) transplantation, viz, low cell dosage and delayed reconstitution, can be overcome by ex vivo expansion. CB expansion under conventional culture causes rapid cell differentiation and depletion of hematopoietic stem and progenitor cells (HSPCs) responsible for engraftment. In this study, we use combinatorial cell culture technology (CombiCult^®) to identify medium formulations that promote CD133⁺ CB HSPC proliferation while maintaining their phenotypic characteristics. We employed second-generation CombiCult screens that use electrospraying technology to encapsulate CB cells in alginate beads. Our results suggest that not only the combination but also the order of addition of individual components has a profound influence on expansion of specific HSPC populations. Top protocols identified by the CombiCult screen were used to culture human CD133⁺ CB HSPCs on nanofiber scaffolds and validate the expansion of the phenotypically defined CD34⁺CD38^lo/-CD45RA^-CD90⁺CD49f⁺ population of hematopoietic stem cells and their differentiation into defined progeny.

SUBMITTER: Tarunina M

PROVIDER: S-EPMC5105341 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Publications

A Novel High-Throughput Screening Platform Reveals an Optimized Cytokine Formulation for Human Hematopoietic Progenitor Cell Expansion.

Tarunina Marina M Hernandez Diana D Kronsteiner-Dobramysl Barbara B Pratt Philip P Watson Thomas T Hua Peng P Gullo Francesca F van der Garde Mark M Zhang Youyi Y Hook Lilian L Choo Yen Y Watt Suzanne M SM

Stem cells and development 20160930 22

The main limitations of hematopoietic cord blood (CB) transplantation, viz, low cell dosage and delayed reconstitution, can be overcome by ex vivo expansion. CB expansion under conventional culture causes rapid cell differentiation and depletion of hematopoietic stem and progenitor cells (HSPCs) responsible for engraftment. In this study, we use combinatorial cell culture technology (CombiCult<sup>®</sup>) to identify medium formulations that promote CD133<sup>+</sup> CB HSPC proliferation while ...[more]

PMID: 27554619

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Project description:Umbilical cord blood (UCB) has been established as an alternative source for hematopoietic stem/progenitor cells (HSPC) for cell and gene therapies. Limited cell yields of UCB units have been tackled with the development of cytokine-based ex vivo expansion platforms. To improve the effectiveness of these platforms, namely targeting clinical approval, in this study, we optimized the cytokine cocktails in two clinically relevant expansion platforms for HSPC, a liquid suspension culture system (CS_HSPC) and a co-culture system with bone marrow derived mesenchymal stromal cells (BM MSC) (CS_HSPC/MSC). Using a methodology based on experimental design, three different cytokines [stem cell factor (SCF), fms-like tyrosine kinase 3 ligand (Flt-3L), and thrombopoietin (TPO)] were studied in both systems during a 7-day culture under serum-free conditions. Proliferation and colony-forming unit assays, as well as immunophenotypic analysis were performed. Five experimental outputs [fold increase (FI) of total nucleated cells (FI TNC), FI of CD34+ cells, FI of erythroid burst-forming unit (BFU-E), FI of colony-forming unit granulocyte-monocyte (CFU-GM), and FI of multilineage colony-forming unit (CFU-Mix)] were followed as target outputs of the optimization model. The novel optimized cocktails determined herein comprised concentrations of 64, 61, and 80 ng/mL (CS_HSPC) and 90, 82, and 77 ng/mL (CS_HSPC/MSC) for SCF, Flt-3L, and TPO, respectively. After cytokine optimization, CS_HSPC and CS_HSPC/MSC were directly compared as platforms. CS_HSPC/MSC outperformed the feeder-free system in 6 of 8 tested experimental measures, displaying superior capability toward increasing the number of hematopoietic cells while maintaining the expression of HSPC markers (i.e., CD34+ and CD34+CD90+) and multilineage differentiation potential. A tailored approach toward optimization has made it possible to individually maximize cytokine contribution in both studied platforms. Consequently, cocktail optimization has successfully led to an increase in the expansion platform performance, while allowing a rational side-by-side comparison among different platforms and enhancing our knowledge on the impact of cytokine supplementation on the HSPC expansion process.

Dataset Information

A Novel High-Throughput Screening Platform Reveals an Optimized Cytokine Formulation for Human Hematopoietic Progenitor Cell Expansion.

Publications

A Novel High-Throughput Screening Platform Reveals an Optimized Cytokine Formulation for Human Hematopoietic Progenitor Cell Expansion.

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