白细胞介素3与c-kit~＋Lin^-细胞的增殖

Effects of interleukin-3 on the proliferation of c-kit+Lin-cells in bone marrow

背景:c-kit+Lin-即为骨髓衍生肝干细胞,与其他造血干细胞类似,c-kit+Lin-细胞的数量有限,难以满足科研的需要。目的:观察在无血清无基质的条件下,白细胞介素3和因子组合对扩增c-kit+Lin-细胞的影响。方法:采用免疫磁珠法分离小鼠骨髓c-kit+Lin-细胞,在干细胞因子、肝细胞生长因子、FLt-3配基、促血小板生成素和白血病抑制因子组合中添加不同剂量的白细胞介素3培养7d,检测细胞总数、c-kit+Lin-细胞扩增倍数及细胞凋亡率。结果与结论:各因子组均可显著扩增c-kit+Lin-细胞,扩增倍数7~19倍不等。随白细胞介素3剂量加大,虽然细胞总数上升,但是超过一定浓度后c-kit+Lin-细胞扩增反而不明显。40μg/L白细胞介素3组细胞总数扩增245.41倍,但c-kit+Lin-细胞扩增仅为15.80倍,与20μg/L白细胞介素3组差异有显著性意义(P<0.05)。但40μg/L白细胞介素3组细胞凋亡率最低,仅为4.66%。提示白细胞介素3能协同干细胞因子+肝细胞生长因子+FLt-3配基+促血小板生成素+白血病抑制因子有效地扩增c-kit+Lin-细胞,其表型并不受影响,但过高剂量反而促使干细胞分化。20μg/L剂量的白细胞介素3扩增效果最佳。

BACKGROUND： c-kit＋Lin- bone marrow derived liver stem cells are similar to other hemopoietic stem cells and not rich in bone marrow, so it is difficult to meet the needs of scientific research. OBJECTIVE： To elucidate the effect of optimal cytokine combinations with interleukin-3 （IL-3） on the proliferation of c-kit＋Lin- cells in bone marrow. METHODS： c-kit＋Lin- cells were isolated from mouse bone marrow by using a high-gradient magnetic cell sorting system （MACS）, and different concentrations of IL-3 were injected into the combinations of stem cell factor （SCF）, hepatic growth factor （HGF）, FLt-3 ligand （FL）, leukemia inhibitor factor （LIF）, thrombopoietin （TPO） and cultured for 7 days in a liquid culture system. The number of cells, multiples of c-kit＋Lin- cell proliferation and the apoptosis rate were detected. RESULTS AND CONCLUSION： Each group could expand c-kit＋Lin- cells rapidly, and the numbers of cells expanded were 7-19 times as large as the beginning’s number. But when the concentration of IL-3 increased and reached to a certain concentration, the c-kit＋Lin- cells did not increased effectively. The expansion of total cells was 245.41 times in 40 μg/L IL-3 group, but the expansion of c-kit＋Lin- cells was only 15.80 times. There was significant difference between the 40 μg/L IL-3 group and 20 μg/L IL-3 group （P 0.05）. And in the 40 μg/L IL-3 group, the apoptosis rate showed the lowest was 4.66%. IL-3 showed a stronger potential for expanding c-kit＋Lin- cells. It showed synergistic actions with SCF, HGF, FL, LIF and TPO to expand c-kit＋Lin- cells and the phenotype was not changed, but too much IL-3 could not increased the c-kit＋Lin- cells effectively. 20 μg/L IL-3 was the best concentration to expand c-kit＋Lin- cells.