Thrombopoietin enhances generation of CD34+ cells from human embryonic stem cells.
The role of thrombopoietin (TPO) in adult hematopoiesis is well-established. A recent report suggests that TPO and vascular endothelial growth factor (VEGF) play a role in promoting formation of early erythropoietic progenitors in a nonhuman primate embryonic stem cell (ES) model. No such report exists for human ES cells as yet. Because TPO may become an important factor promoting human ES cell-derived hematopoiesis, we sought to investigate whether TPO in combination with VEGF can enhance human ES-derived hematopoiesis in an EB-derived culture system. The emphasis of this work was to demonstrate the molecular mechanisms involved in this process, specifically the role of c-mpl and its ligand TPO. Human ES cells were cultured to the EB state, and EB-derived secondary cultures supporting hematopoietic differentiation were established: condition 1, control (stem cell factor [SCF] and Flt3 ligand [Flt3L]); condition 2, SCF, Flt3L, and TPO; and condition 3, SCF, Flt3L, TPO, and VEGF. Cells were harvested daily, starting at day 2 and continuing until day 8, for reverse transcription-polymerase chain reaction and Western blot. There was no evidence of expression of c-mpl and VEGF receptor on the gene or protein level until day 8, when the formation of well-established hematopoietic colonies began. This correlated with the formation of CD34+/CD31- negative progenitors, mostly found in blast-forming units-erythroid-like colonies. We concluded that TPO and VEGF play an important synergistic role in the formation of early ES-derived hematopoietic progenitors that occurs through the c-mpl and VEGF receptors. Disclosure of potential conflicts of interest is found at the end of this article.
Srivastava
Anand S
AS
Department of Medicine, Pediatrics and UCSD Moores Cancer Center, School of Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
Nedelcu
Elena
E
Esmaeli-Azad
Babak
B
Mishra
Rangnath
R
Carrier
Ewa
E
D018952:Antigens, CD34 / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D053628:Receptors, Thrombopoietin / Q000235:genetics / Q000378:metabolism; D040262:Receptors, Vascular Endothelial Growth Factor / Q000235:genetics / Q000378:metabolism; D013926:Thrombopoietin / Q000494:pharmacology* / Q000502:physiology
D018952:Antigens, CD34; D053628:Receptors, Thrombopoietin; C082218:MPL protein, human; D013926:Thrombopoietin; D040262:Receptors, Vascular Endothelial Growth Factor