[Effects of recombinant human macrophage colony stimulating factor (rhM-CSF) on stromal cell derived during the time of mice colony formation unit-spleen (CFU-S) and its role on CD34+ cells expansion in vitro].

OBJECTIVE

To investigate the effects of stromal cell derived from mice spleen during in vitro expansion of CD34+ hematopoietic stem cells from umbilical cord blood.

METHODS

Irradiated and non-irradiated mice at day 10, 12 and 14 were sacrificed and their spleens were taken for collecting spleen cells. The cells were stimulated with recombinant human macrophage colony stimulating factor(rhM-CSF) for collecting adherent spleen derived stroma cells. The effects of spleen derived stromal cells which treated with mitomycin served as a feeder layer, combined with different cytokines on in vitro expansion of purified cord blood CD34+ were assayed. Clonogenic assay and flow cytometry were employed to analyze phenotype of expanded cells and their ability to form different colony in vitro.

RESULTS

1. rhM-CSF efficiently stimulated spleen derived stromal cells to proliferate; 2) The mice stromal cells obtained during the time of colony formation-unit spleen (CFU-S) promoted the CD34+ cells to form colony in semisolid medium and, combined with cytokines, thereby efficiently expanded the hematopoietic cells from cord blood. After two weeks incubation, CD34 positive cells were 100 times more than original cells cultured and expanded cells still have the ability to form multi-lineage colony in vitro.

CONCLUSION

rhM-CSF can stimulate the stromal cells derived from CFU-S to proliferate and support the survival and expansion of CD34+ cells separated from cord blood.