目的 探讨骨髓造血细胞的冷冻保存效果及人参总皂苷 (TSPG)降低冷冻损伤的作用。方法 以 5 %二甲基亚砜 (DMSO)为冷冻保护剂,采用逐步降温法将骨髓造血细胞在液氮中保存 3～ 5个月,复苏后应用台盼蓝拒染法、集落形成试验、CD34+及细胞总数回收率等,测定骨髓造血细胞生物学活性的变化 ;将不同剂量的 TSPG与复苏的骨髓造血细胞共培养,应用集落形成试验、MTT比色分析法及流式细胞术等方法检测 TSPG对骨髓造血细胞冷冻损伤可恢复性的影响。结果 骨髓造血细胞在低温保存时一部分细胞会失去增殖能力,但这种损伤并不与保存时间成正比。细胞复苏后,加入合适剂量的 TSPG(2 5 μg/ m L),可明显提高冻存骨髓造血细胞的集落产率 ;TSPG在一定浓度范围内 (2 5～ 5 0μg/ m L )能促进冻存骨髓造血细胞的增殖 ;TSPG (2 5μg/ m L )能促进冻存骨髓造血细胞进入细胞增殖周期。结论 TSPG能促进冷冻骨髓造血细胞的复苏、增殖,提高骨髓细胞冷冻损伤的可恢复性,对应用冻存骨髓造血细胞进行造血移植有一定意义。

Object To study the effect of cryopreservation on the ability of hematopoietic stem cells from human bone marrow and the action of total saponins of Panax ginseng (TSPG) to reduce their freezing injury. Methods Gradual cooling methods were used to place mononuclear cells (MNC) in liquidnitrogen with 5% dimethylsulfoxide (DMSO) for 3—5 months. After thawed, the biological properties of MNC were monitored, which included the mean trypan blue exclusion rate and the mean recovery rate of MNC, CFU-Mix, CD34⁺ cell, and total cells; Thawed MNC were cultured with various concentrations of TSPG, the effect of TSPG on the recoverable ability from cry opreservation damage were detected by colony-forming assay, colorimetric MTT assay for cell proliferation and f low cy tometry. Results A fraction of MC lost their proliferatory ability after thaw ed, but the damage was not deteriorated with freezing time. TSPG (25μg/mL) could raise the colony production rate of thawed hemato poietic stem cells;TSPG (25—50μg/mL) could raise the colony production rate of thawed hematopoietic stemcells; TSPG(25—50μg/mL) could improve their proliferation; TSPG (25 μg /mL) could also promote the entray of them into cell prolifera to ry cycle. Conclusion TSPG could induce the proliferatio n of thaw ed hema topoietic stem cells and raise the post-freezing recoverable ability of hemato poietic stem cells.

国家自然科学基金资助项目(38670880);重庆市科委重点攻关课题(渝科委计[2000]25)