NK cells differentiated from bone marrow, cord blood and peripheral blood stem cells exhibit similar phenotype and functions

Abstract

In the present study, we investigated the differentiation of human NK cells from bone marrow, cord blood and mobilized peripheral blood purified CD34+ stem cells using a potent culture system. Elutriated CD34+ stem cells were grown for several weeks in medium supplemented with stem cell factor (SCF) and IL-15 in the presence or absence of a murine stromal cell line (MS-5). Our data indicate that IL-15 induced the proliferation and maturation of highly positive CD56+ NK cells in both types of culture, although murine stromal cells slightly increased the proliferation of NK cells. NK cells differentiated in the presence of MS-5 were mostly CD56+ CD7 and a small subset expressed CD16. These in vitro differentiated CD56+ NK cells displayed cytolytic activity against the HLA class I- target K562. The CD56+ CD16+ subset also lysed NK-resistant Daudi cells. Neither of these NK subsets were shown to express Fas ligand. Total CD56+ cells expressed high amounts of transforming growth factor-beta and granulocyte-macrophage colony-stimulating factor, but no IFN-gamma. Investigation of NK receptor expression showed that most CD56+ cells expressed membrane CD94 and NKG2-A mRNA. PCR analysis revealed that p58 was also expressed in these cells. The role of CD94 in NK cell-mediated cytotoxicity was assessed on human HLA-B7-transfected murine L cells. While a low cytotoxic activity towards HLA-B7 cells was observed, the HLA-DR4 control cells were killed with high efficiency. These studies demonstrate that cytolytic and cytokine-producing NK cells may be derived from adult and fetal precursors by IL-15 and that these cells express a CD94 receptor which may influence their lytic potential.

Altered natural killer cell differentiation in CD34+ progenitors from Chronic Myeloid Leukemia patients

“…Functionally, CD56 bright cells lysed K562 targets, although with a lower e ciency than NK cells derived from normal progenitors, whereas CD56 low and CD56 7 cells did not exert any cytotoxic activity against K562 (Figure 1) or p46 redirected of P815 (data not shown). These data imply that the lytic potential of these e ectors clearly correlate with the intensity of their CD56 expression, further con®rming our previous observation (Carayol et al, 1998).…”

“…Immunoselected leukemic CD34 + cells were cultured in the presence of SCF and IL-15, as previously described (Carayol et al, 1998). The results obtained from the 20 patients are summarized in Table 1.…”

“…Mononuclear cells were isolated after centrifugation on Ficoll gradient (Lymphoprep Nicomed Pharma SA, Oslo, Norway). Selection of CD34 + cells was performed by immunomagnetic selection (Miltenyi, Tebu, France) on mononuclear cells labeled with a mAb speci®c for the QBEND10 epitope of the CD34 antigen as previously described (Carayol et al, 1998 In vitro culture of CD34 + cells…”

“…Di erentiated NK cells were tested for cytotoxicity against the NK sensitive cell line K562 in a 4 h Cr 51 release assay (Carayol et al, 1998). E ector to target ratios ranged from 10 : 1 to 2 : 1.…”

“…For this purpose we used a previously described in vitro system to di erentiate NK cells from CD34 + stem cells using combination of SCF and IL-15 (Carayol et al, 1998). IL-15 is produced by bone marrow stromal cells and recently, it was found as the most powerful cytokine to drive CD34 + cell di erentiation into NK cells (Carson et al, 1994(Carson et al, , 1997Mingari et al, 1997).…”