Long-term platelet production assessed in NOD/SCID mice injected with cord blood CD34+ cells, thrombopoietin-amplified in clinical grade serum-free culture.
OBJECTIVE
Delayed platelet recovery post-cord blood (CB) transplantation might be due to CB characteristics: low maturity of stem cell compartment, poor production of CD34+/CD41+ cells when induced to differentiate along the megakaryocytic (MK) lineage, retention of a low ploidy in the expanded MKs. Ex vivo expansion of CB hematopoietic progenitor cells for reconstitution of different human hematopoietic lineages has already been developed in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. However, optimal conditions for MK-progenitor engraftment to reduce hemorrhaging risk still to be developed. This study assesses the hypothesis that CB-CD34+ amplification with thrombopoietin (TPO) can be applied to a portion of a CB transplant unit to stimulate recovery along MK differentiation program.


METHODS
Human CB-CD34+ cells were amplified in a serum-free, clinical grade medium with 100 ng/mL TPO alone and in addition to other cytokines (Kit ligand, interleukin-6, and Flt-3 ligand). Seven-day cultured cells were transplanted into irradiated NOD/SCID mice and engraftment, megakaryocytopoiesis, and platelet production were assessed.


RESULTS
Platelet release was successful and continuously present for at least 8 weeks in NOD/SCID mice transplanted with CB cells stimulated by TPO. Thrombocytopoiesis was more effective with transplanted TPO-amplified cells than with the cytokine cocktails.


CONCLUSIONS
Platelet number obtained is within the minimum level considered sufficient for hemostasis. Furthermore, amplified cells maintain their self-renewal capacity and multilineage potential differentiation. Thus, transplantation of TPO-expanded CB cells has the potential favoring both platelet recovery and human engraftment.
Mattia
Gianfranco
G
Department of Hematology, Oncology and Molecular medicine, Section of Transfusion Methodologies, Istituto Superiore di Sanità, Rome, Italy.
Milazzo
Luisa
L
Vulcano
Francesca
F
Pascuccio
Massimiliano
M
Macioce
Giampiero
G
Hassan
Hamisa Jane
HJ
Giampaolo
Adele
A
D000818:Animals; D018952:Antigens, CD34; D001792:Blood Platelets / Q000166:cytology / Q000502:physiology*; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D036101:Cord Blood Stem Cell Transplantation; D016895:Culture Media, Serum-Free; D005312:Fetal Blood / Q000166:cytology / Q000502:physiology*; D006085:Graft Survival / Q000187:drug effects / Q000502:physiology; D006470:Hemorrhage / Q000378:metabolism / Q000517:prevention & control; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology; D008533:Megakaryocytes / Q000166:cytology / Q000502:physiology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D012307:Risk Factors; D019089:Stem Cell Factor / Q000494:pharmacology; D034061:Thrombopoiesis / Q000187:drug effects* / Q000502:physiology; D013926:Thrombopoietin / Q000494:pharmacology*; D013997:Time Factors; D014183:Transplantation, Heterologous; D014184:Transplantation, Homologous
D018952:Antigens, CD34; D016895:Culture Media, Serum-Free; C508600:IL6 protein, human; D015850:Interleukin-6; D019089:Stem Cell Factor; D013926:Thrombopoietin