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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- is a bone marrow-derived hepatic stem cell. Similar to other hematopoietic stem cells, the number of c-kit + Lin- cells is limited and can not meet the needs of scientific research. OBJECTIVE: To investigate the effects of interleukin-3 and factor combination on the expansion of c-kit + Lin- cells in serum-free and stromal-free conditions. Methods: The mouse bone marrow c-kit + Lin- cells were isolated by immunomagnetic beads method and different doses of leukocyte mediators were added to the combination of stem cell factor, hepatocyte growth factor, FLt-3 ligand, thrombopoietin and leukemia inhibitory factor The cells were cultured for 7 days. The total number of cells, c-kit + Lin-cell multiplication and apoptosis rate were detected. RESULTS AND CONCLUSION: C-kit + Lin- cells were significantly expanded in all factor groups with multiples of 7-19 fold. With the increase of interleukin-3 dose, although the total number of cells increased, c-kit + Lin- cells expanded more than a certain concentration but not obvious. The total number of cells in 40μg / L interleukin-3 group was 245.41 times of that in c-kit + Lin-cells, but only 15.80 times that of 20μg / L interleukin-3 group (P <0.05). However, the apoptosis rate of 40μg / L interleukin 3 group was the lowest, only 4.66%. It is suggested that interleukin-3 can effectively expand c-kit + Lin- cells in combination with stem cell factor + hepatocyte growth factor + FLt-3 ligand + thrombopoietin + leukemia inhibitory factor and its phenotype is unaffected Excessively high doses, on the contrary, promote stem cell differentiation. 20μg / L dose of interleukin 3 amplification best.