Presence of primitive lymphoid progenitors with NK or B potential in ex vivo expanded bone marrow cell cultures.
OBJECTIVE
In previous work, we showed that CD34+ bone marrow cells can be successfully expanded along the myeloid pathway in stroma- and serum-free conditions in the presence of SCF+IL-3+IL-6+Flt3-l+G-CSF+MGDF. Due to the lack of phenotypically detectable lymphoid cells, it was necessary to address the question of the lymphoid potential of the expanded populations under these conditions.


METHODS
The present report describes a long-term culture system that supports human B- and NK-cell differentiation from the day 14 fraction without further selection of the more primitive cells. In NK proliferation assays, the cells were maintained over stroma cells in the presence of IL-2 for 4-5 weeks. NK initiating cells (NK-IC) were determined by a limiting dilution assay. In B-cell cultures, the expanded cells were maintained over MS5 in the presence of Flt3-l for 4-8 weeks.


RESULTS
NK cells rose from 0.2%+/-0.04% at culture initiation to 71%+/-6% at week 5. These cells displayed cytolytic activity. NK-IC evaluation showed a mean 18-fold expansion in the day 14 expanded fraction as compared to the initial day 0 fraction. Similarly, CD19+ cells rose from 0.1% at culture initiation to 30%+/-1% at week 6. Cells produced under these B-LTC conditions were CD34-CD19+CD10+. We also demonstrated that the CD34+/Lin- sorted cells from the day 14 fraction gave rise to NK and B cells.


CONCLUSIONS
This culture system permits the revelation of a population that, although poorly represented in terms of phenotypically detectable cells, nevertheless retains high levels of lymphoid NK and B potential after 14 days expansion. Such data suggest the persistence, or expansion, of lymphoid progenitors and, hence, the multipotentiality of the expanded progenitor/stem cells.
Giarratana
M C
MC
INSERM U417, Hôpital Saint-Antoine, Paris.
Vergé
V
V
Schmitt
C
C
Bertho
J M
JM
Kobari
L
L
Barret
C
C
Douay
L
L
D018952:Antigens, CD34 / Q000096:biosynthesis; D001402:B-Lymphocytes / Q000166:cytology*; D001854:Bone Marrow Cells / Q000166:cytology*; D017252:CD3 Complex / Q000096:biosynthesis; D019002:CD56 Antigen / Q000096:biosynthesis; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002455:Cell Division; D002478:Cells, Cultured; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007694:Killer Cells, Natural / Q000166:cytology* / Q000378:metabolism; D008214:Lymphocytes / Q000166:cytology* / Q000378:metabolism
D018952:Antigens, CD34; D017252:CD3 Complex; D019002:CD56 Antigen