Cycling status of CD34+ cells mobilized into peripheral blood of healthy donors by recombinant human granulocyte colony-stimulating factor

R M Lemoli, A Tafuri, A Fortuna, M T Petrucci, M R Ricciardi, L Catani, D Rondelli, M Fogli, G Leopardi, C Ariola, S Tura
Blood 1997 February 15, 89 (4): 1189-96
In this study, we assessed the functional and kinetic characteristics of highly purified hematopoietic CD34+ cells from the apheresis products of 16 normal donors undergoing glycosylated granulocyte colony-stimulating factor (G-CSF) treatment for peripheral blood stem cells (PBSC) mobilization and transplantation in allogeneic recipients. Mobilized CD34+ cells were evaluated for their colony-forming capacity and trilineage proliferative response to selected recombinant human (rh) CSF in vitro and the content of very primitive long-term culture initiating cells (LTC-IC). In addition, the cycling status of circulating CD34+ cells, including committed clonogenic progenitor cells and the more immature LTC-IC, was determined by the cytosine arabinoside (Ara-C) suicide test and the acridine orange flow cytometric technique. By comparison, bone marrow (BM) CD34+ cells from the same individuals were studied under steady-state conditions and during G-CSF administration. Clonogenic assays in methylcellulose showed the same frequency of colony-forming unit cells (CFU-C) when PB-primed CD34+ cells and BM cells were stimulated with phytohemagglutinin-lymphocyte-conditioned medium (PHA-LCM). However, mobilized CD34+ cells were significantly more responsive than their steady-state BM counterparts to interleukin-3 (IL-3) and stem cell factor (SCF) combined with G-CSF or IL-3 in presence of erythropoietin (Epo). In cultures added with SCF, IL-3, and Epo, we found a mean increase of 1.5- +/- 1-fold (standard error of the mean [SEM]) of PB CFU-granulocyte-macrophage and erythroid progenitors (burst-forming units-erythroid) as compared with BM CD34+ cells (P < .05). Conversely, circulating and BM megakaryocyte precursors (CFU-megakaryocyte) showed the same clonogenic efficiency in response to IL-3, granulocyte-macrophage-CSF and IL-3, IL-6, and Epo. After 5 weeks of liquid culture supported by the engineered murine stromal cell line M2-10B4 to produce G-CSF and IL-3, we reported 48.2 +/- 35 (SEM) and 62.5 +/- 54 (SEM) LTC-IC per 10(4) CD34+ cells in PB and steady-state BM, respectively (P = not significant). The Ara-C suicide assay showed that 4% +/- 5% (standard deviation [SD]) of committed precursors and 1% +/- 3% (SEM) of LTC-IC in PB are in S-phase as compared with 25.5% +/- 12% (SD) and 21% +/- 8% (SEM) of baseline BM, respectively (P < .001). However, longer incubation with Ara-C (16 to 18 hours), in the presence of SCF, IL-3 and G-CSF, or IL-6, showed that more than 60% of LTC-IC are actually cycling, with no difference being found with BM cells. Furthermore, studies of cell-cycle distribution on PB and BM CD34+ cells confirmed the low number of circulating progenitor cells in S- and G2M-phase, whereas simultaneous DNA/RNA analysis showed that the majority of PB CD34+ cells are not quiescent (ie, in G0-phase), being in G1-phase with a significant difference with baseline and G-CSF-treated BM (80% +/- 5% [SEM] v 61.9% +/- 6% [SEM] and 48% +/- 4% [SEM], respectively; P < .05). Moreover, G-CSF administration prevented apoptosis in a small but significant proportion of mobilized CD34+ cells. Thus, our results indicate that mobilized and BM CD34+ cells can be considered equivalent for the frequency of both committed and more immature hematopoietic progenitor cells, although they show different kinetic and functional profiles. In contrast with previous reports, we found that PB CD34+ cells, including very primitive LTC-IC, are cycling and ready to progress into S-phase under CSF stimulation. This finding should be taken into account for a better understanding of PBSC transplantation.

Full Text Links

Find Full Text Links for this Article


You are not logged in. Sign Up or Log In to join the discussion.

Trending Papers

Remove bar
Read by QxMD icon Read

Save your favorite articles in one place with a free QxMD account.


Search Tips

Use Boolean operators: AND/OR

diabetic AND foot
diabetes OR diabetic

Exclude a word using the 'minus' sign

Virchow -triad

Use Parentheses

water AND (cup OR glass)

Add an asterisk (*) at end of a word to include word stems

Neuro* will search for Neurology, Neuroscientist, Neurological, and so on

Use quotes to search for an exact phrase

"primary prevention of cancer"
(heart or cardiac or cardio*) AND arrest -"American Heart Association"