Cell Cycle-Related Changes in Repopulating Capacity of Human Mobilized Peripheral Blood CD34+ Cells in Non-Obese Diabetic/Severe Combined Immune-Deficient Mice

Abstract

Most primitive hematopoietic progenitor cells reside in vivo within the G0/G1 phase of the cell cycle. By simultaneous DNA/RNA staining it is possible to distinguish G0 and G1 states and to isolate cells in defined phases of the cell cycle. We report here the use of cell cycle fractionation to separate human mobilized peripheral blood (MPB) CD34+ cells capable of repopulating the bone marrow (BM) of non-obese diabetic/severe combined immune-deficient (NOD/SCID) mice. In freshly isolated MPB, repopulating cells were predominant within the G0 phase, because transplantation of CD34+cells residing in G0 (G0CD34+) resulted on average in a 16.6- ± 3.2-fold higher BM chimerism than infusion of equal numbers of CD34+ cells isolated in G1. We then investigated the effect of ex vivo cell cycle progression, in the absence of cell division, on engraftment capacity. Freshly isolated G0CD34+ cells were activated by interleukin-3 (IL-3), stem cell factor (SCF), and flt3-ligand (FL) for a 36-hour incubation period during which a fraction of cells progressed from G0 into G1 but did not complete a cell cycle. The repopulating capacity of stimulated cells was markedly diminished compared with that of unmanipulated G0CD34+ cells. Cells that remained in G0 during the 36-hour incubation period and those that traversed into G1 were sorted and assayed separately in NOD/SCID recipients. The repopulating ability of cells remaining in G0 was insignificantly reduced compared with that of unstimulated G0CD34+ cells. On the contrary, CD34+ cells traversing from G0 into G1 were largely depleted of repopulating capacity. Similar results were obtained when G0CD34+ cells were activated by the combination of thrombopoietin-SCF-FL. These studies provide direct evidence of the quiescent nature of cells capable of repopulating the BM of NOD/SCID mice. Furthermore, these data also demonstrate that G0-G1 progression in vitro is associated with a decrease in engraftment capacity. © 1998 by The American Society of Hematology.

Functional and kinetic characterization of granulocyte colony‐stimulating factor‐primed CD34− human stem cells

“…*P-value < 0AE05. capacity of human mobilized PB CD34 + cells in NOD/SCID mice was shown to be mainly expressed in G 0 cells as opposed to G 1 cells (Gothot et al, 1998). This is in contrast with the kinetic status and the repopulating capacity of CD34 + and CD34 ) cells shown in this study.…”

The Late Dividing Population of γ‐Retroviral Vector Transduced Human Mobilized Peripheral Blood Progenitor Cells Contributes Most to Gene‐Marked Cell Engraftment in Nonobese Diabetic/Severe Combined Immunodeficient Mice

“…In response to cytokines, the majority of cells go into cell cycle. Compared with CD34ϩ cells in G 0 , cells in cell cycle phase G 1 , and even more so, those in S/G 2 phase, demonstrate diminished engraftment potential [7][8][9][10][11]. This is consistent with and equivalent to the observation that rapidly cycling cells are shown to be committed progenitors [5,6].…”

Different behaviour of fresh and cultured CD34+ cells during immunomagnetic separation

“…In addition, impaired engraftment of cultured cells has been reported (Brandt et al, 1998;Habibian et al, 1998;Gothot et al, 1998), which may also be due to changes in cell surface antigens induced by in vitro culture. These changes may be a reversible plastic feature correlated with cell cycle progression (Habibian et al, 1998), including decreased expression of adhesion proteins during late S/G 2 (Becker et al, 1997) or an increase in peptides which allow cells to undergo endothelial transmigration so that infusion of cells in cycle would lead to the homing of cells in non-haemopoietic organs (Yong et al, 1998).…”