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Paper Information

Journal:   CELL JOURNAL (YAKHTEH)   FALL 2009 , Volume 11 , Number SUPPL. 1; Page(s) 104 To 105.
 
Paper: 

PLATELET GROWTH FACTORS SUPPRESS THE EXPANSION, BUT PROMOTE THE DIFFERENTIATION OF CORD BLOOD CD133+ STEM CELLS TO MEGAKARYOCYTE PROGENITORS

 
 
Author(s):  NOUROUZI AGHIDEH A.*, KHEYR ANDISH MARYAM, ABOU ALGHASEMI HASAN, GHARAH BAGHIAN A., KHALAF ADELI E., SIADAT S.D., HAGHBIN S.
 
* RESEARCH CENTER, IRANIAN BLOOD TRANSFUSION ORGANIZATION (IBTO), TEHRAN, IRAN
 
Abstract: 

Objective: Umbilical cord blood (UCB) is a rich source of hematopoietic cells as a valuable source for hematopoietic stem cell transplantation (HSCT). In addition, cord blood megakaryocytic progenitor cells are more immature than other sources such as bone marrow (BM) that result in delayed platelet recovery in the recipients of UCB transplants. Platelet rich plasma (PRP) is a concentrate of platelet growth factors and many studies have shown the effects of these factors on the expansion and function of different cell types. In this study, we surveyed the effects of platelet growth factors including the platelet supernatant (PS), platelet lysate (PL) and activated PRP (AP) on the expansion and differentiation of cord blood CD133+ stem cells into megakaryocytic progenitor cells.
Materials and Methods: UCB 133+ cells were separated by MACS method and counted using improved neubauer hemocytometer their viability was determined using dye exclusion assay and 7-AAD method. AP was prepared by PRP activation by thrombin and calcium chloride. PS and PL preparation was performed by high speed centrifugation (900 g) of PRP and freezing and thawing of PRP, respectively. The growth factors and protein concentrations in these platelet products were measured by ELISA and Bradford methods, respectively. Mean expression rate of CD133, CD41, CD61 and CD42b in day 0 and CD41, CD61 and CD42b at the end of culture period were assayed by flow cytometry.
Results: The results showed that PDGF and TGF-
b concentrations are higher than other platelet growth factors. Further, protein concentrations in PS, PL and AP were 460, 490 and 480 mg/ml, respectively (p<0.05). AP at 2, 10, 25, 50 mg/ml and PS at 50 and 100 mg/ml concentrations significantly suppressed the expansion of CD133+ cells in the first day of the culture (p <0.05). This suppression for PL at 50mg/ml and for AP at 10, 25 and 50 mg/ml concentrations after the fourth day of culture was not significant (p>0.05). On the other hand, the expression of CD41, CD61 and CD42b markers in the presence of all growth factors increased compared with the control media, but only AP at the 10, 25, 50 mg/ml concentrations in the first day of culture showed a significant effect on the expression of CD41 and CD61 markers (p<0.05). However, increasing in the expression of CD42b as a late marker of megakaryocytic lineage was not significant under none of the above conditions.
Conclusion: Taken together, platelet growth factors suppressed the expansion of UCB CD133+ cells and increased the differentiation of UCB CD133+ cells into megakaryocytic progenitor cells in a dose and time dependent manner. In our study, amplifying of UCB CD133+ cells by cytokine cocktail and then promotion of their differentiation using platelet growth factors appeared to be an efficient strategy to overcome the low numbers in UCB units that limits their use in full reconstitution of adult BM.

 
Keyword(s): UMBILICAL CORD BLOOD, CD133+ CELLS, MEGAKARYOCYTE PROGENITORS, EXPANSION, DIFFERENTIATION, PLATELET GROWTH FACTORS
 
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