Abstract
Background aims. Ex vivo generation of megakaryocytes (MK) from hematopoietic stem cells (HSC) is important for both basic research, to understand the mechanism of platelet biogenesis, and clinical infusions, for rapid platelet recovery in thrombocytopenic patients. We investigated the role of two nutraceuticals, docosahexanoic acid (DHA) and arachidonic acid (AA), in the in vitro generation of MK. Methods. Umbilical cord blood (UCB)-derived CD34+cells were cultured with stem cell factor (SCF) and thrombopoietin (TPO) in the presence (test) or absence (control) of the two additives. On day 10, MK and platelets generated were quantitated by morphologic, phenotypic and functional assays. Results. The cell yield of MK and platelet numbers were significantly higher in test compared with control cells. Phenotypic analyzes and gene expression profiles confirmed these findings. Functional properties, such as colony-forming unit (CFU)-MK formation, chemotaxis and platelet activation, were found to be enhanced in cells cultured with nutraceuticals. The engraftment potential of ex vivo-expanded cells was studied in NOD/SCID mice. Mice that received MK cultured in the presence of DHA/AA engrafted better. There was a reduction in apoptosis and total reactive oxygen species (ROS) levels in the CD41+ compartment of the test compared with control sets. The data suggest that these compounds probably exert their beneficial effect by modulating apoptotic and redox pathways. Conclusions. Use of nutraceuticals like DHA and AA may prove to be a useful strategy for efficient generation of MK and platelets from cord blood cells, for future use in clinics and basic research.
Acknowledgements
The authors thank Drs Prakash Daithankar, Arvind Sangamnerkar Girish Godbole and Charulata Bapaye for providing cord blood samples and Arvind Marathe for helping in the identification of cell types. This work was supported by a grant from the Department of Biotechnology (DBT), Government of India. NS received Assistantship from DBT.
Statement of equal author's contribution
NFS and NS contributed equally to this manuscript. Parts of this manuscript were presented as posters by LSL at the 35th Annual Scientific Meeting of the International Society for Experimental Hematology, held in September 2006, at Minneapolis, USA, and by NFS at the International Conference on Stem Cell Research held at Bangalore, India, in January 2007.
Authorship and disclosures
The authors reported no potential conflicts of interest.