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Mesenchymal stem cells promote a primitive phenotype CD34+c-kit+ in human cord blood-derived hematopoietic stem cells during ex vivo expansion

Abstract

The purpose of this study was to evaluate the influence of bone marrow-mesenchymal stem cells (BM-MSC) and exogenously added cytokines on the proliferation, primitive cell subpopulation maintenance (including the c-kit+ marker) and clonogenic capacity of hematopoietic stem cells (HSC). BM-MSC were collected from volunteer donors, isolated and characterized. Umbilical cord blood (UCB) samples were collected from healthy full-term deliveries. UCB-CD34+ cells were cultured in the presence or absence of BM-MSC and/or cytokines for 3 and 7 days. CD34+ cell proliferation was evaluated using the CSFE method and cell phenotype was determined by CD34, c-kit, CD33, CD38, HLA-DR, cyCD22 and cyCD3 detection. Cell clonogenic ability was also assessed. Exogenously added SCF, TPO and FLT3L increasedCD34+ cell proliferation in the presence or absence of BM-MSC, but with concomitant cell differentiation. Without any added cytokines, BM-MSC are able to increase the percentage of primitive progenitors as evaluated by c-kit expression and CFU-GEMM increase. Interestingly, this latter effect was dependent on both cell-cell interactions and secreted factors. A 7-day co-culture period will be optimal for obtaining an increased primitive HSC level. Including c-kit as a marker for primitive phenotype evaluation has shown the relevance of BM-MSC and their secreted factors on UCB-HSC stemness function. This effect could be dissociated from that of the addition of exogenous cytokines, which induced cellular differentiation instead.

Abbreviations

BFU-E:

burst-forming unit erythroids

BM:

bone marrow

BMMSC:

bone marrow mesenchymal stem cells

CFSE:

carboxyfluorescein diacetate succinimidyl ester

CFU-E:

colony-forming unit erythroids

CFU-GEMM:

colonyforming unit-granulocyte, erythrocyte, monocyte, megakaryocyte

CFU-GM:

colonyforming unit-granulocyte, monocyte, macrophage

Flt-3L:

FMS-like tyrosine kinase 3 ligand

FRS:

frozen supernatant

FS:

fresh supernatant

HC:

hematopoietic cytokines

HM:

hematopoietic mesenchymal

HMC:

hematopoietic-mesenchymal cytokines

HSC:

hematopoietic stem cells

LTC-IC:

long-term culture-initiating cell

MACS:

magneticactivated cell sorting

MFI:

medium fluorescence intensity

MSC:

mesenchymal stem cells

SCF:

stem cell factor

TPO:

thrombopoietin

UCB:

umbilical cord blood

UCBCD34+:

umbilical cord blood-hematopoietic stem cell

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Rodríguez-Pardo, V.M., Vernot, J.P. Mesenchymal stem cells promote a primitive phenotype CD34+c-kit+ in human cord blood-derived hematopoietic stem cells during ex vivo expansion. Cell Mol Biol Lett 18, 11–33 (2013). https://doi.org/10.2478/s11658-012-0036-1

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Key words

  • Hematopoietic stem cell
  • Mesenchymal stem cell
  • Bone marrow stroma
  • CD34+c-kit+ subpopulation
  • Cytokines
  • Progenitor cell
  • Umbilical cord blood