Differential ability of MSCs isolated from placenta and cord as feeders for supporting ex vivo expansion of umbilical cord blood derived CD34(+) cells.
BACKGROUND
Ex vivo expansion of umbilical cord blood (UCB) is attempted to increase cell numbers to overcome the limitation of cell dose. Presently, suspension cultures or feeder mediated co-cultures are performed for expansion of hematopoietic stem cells (HSCs). Mesenchymal stem cells (MSCs) have proved to be efficient feeders for the maintenance of HSCs. Here, we have established MSCs-HSCs co-culture system with MSCs isolated from less invasive and ethically acceptable sources like umbilical cord tissue (C-MSCs) and placenta (P-MSCs). MSCs derived from these tissues are often compared with bone marrow derived MSCs (BM-MSCs) which are considered as a gold standard. However, so far none of the studies have directly compared C-MSCs with P-MSCs as feeders for ex vivo expansion of HSCs. Thus, we for the first time performed a systematic comparison of hematopoietic supportive capability of C and P-MSCs using paired samples.


METHODS
UCB-derived CD34(+) cells were isolated and co-cultured on irradiated C and P-MSCs for 10 days. C-MSCs and P-MSCs were isolated from the same donor. The cultures comprised of serum-free medium supplemented with 25 ng/ml each of SCF, TPO, Flt-3 L and IL-6. After 10 days cells were collected and analyzed for phenotype and functionality.


RESULTS
C-MSCs and P-MSCs were found to be morphologically and phenotypically similar but exhibited differential ability to support ex vivo hematopoiesis. Cells expanded on P-MSCs showed higher percentage of primitive cells (CD34(+)CD38(-)), CFU (Colony forming unit) content and LTC-IC (Long term culture initiating cells) ability. CD34(+) cells expanded on P-MSCs also exhibited better in vitro adhesion to fibronectin and migration towards SDF-1α and enhanced NOD/SCID repopulation ability, as compared to those grown on C-MSCs. P-MSCs were found to be closer to BM-MSCs in their ability to expand HSCs. P-MSCs supported expansion of functionally superior HSCs by virtue of reduction in apoptosis of primitive HSCs, higher Wnt and Notch activity, HGF secretion and cell-cell contact. On the other hand, C-MSCs facilitated expansion of progenitors (CD34(+)CD38(+)) and differentiated (CD34(-)CD38(+)) cells by secretion of IL1-α, β, MCP-2, 3 and MIP-3α.


CONCLUSIONS
P-MSCs were found to be better feeders for ex vivo maintenance of primitive HSCs with higher engraftment potential than the cells expanded with C-MSCs as feeders.
Stem Cell Laboratory, National Centre for Cell Science, University of Pune Campus, Ganeshkhind, Pune, 411007, Maharashtra, India. darsh311@gmail.com.
Darshana
D
Kadekar
Stem Cell Laboratory, National Centre for Cell Science, University of Pune Campus, Ganeshkhind, Pune, 411007, Maharashtra, India. vpkale@nccs.res.in.
Vaijayanti
V
Kale
Stem Cell Laboratory, National Centre for Cell Science, University of Pune Campus, Ganeshkhind, Pune, 411007, Maharashtra, India. lslimaye@nccs.res.in.
Lalita
L
Limaye
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D002450:Cell Communication; D002454:Cell Differentiation; D049109:Cell Proliferation; D018920:Coculture Techniques; D061252:Feeder Cells / Q000502:physiology; D005260:Female; D005312:Fetal Blood / Q000166:cytology; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000502:physiology; D006801:Humans; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D010920:Placenta / Q000166:cytology; D011247:Pregnancy; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011833:Radiation Injuries, Experimental / Q000628:therapy; D051880:Receptors, Notch / Q000378:metabolism; D015398:Signal Transduction
D018952:Antigens, CD34; C000595707:BCL2 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D051880:Receptors, Notch