High-efficiency recovery of functional hematopoietic progenitor and stem cells from human cord blood cryopreserved for 15 years
High-efficiency recovery of functional hematopoietic progenitor and stem cells from human cord blood cryopreserved for 15 years
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DOI：
10.1073/pnas.0237086100
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发表时间：
2003-01-21
影响因子：
11.1
通讯作者：
Bodine, DM
中科院分区：
文献类型：
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作者：
Broxmeyer, HE;Srour, EF;Bodine, DM
移植脐带血造血干细胞（HSC）和祖细胞（HPC）可以治疗恶性和非恶性疾病。由于长期冷冻保存对C8细胞库和移植至关重要，我们评估了从冷冻保存15年的单个CB中回收活HSC/HPC的效率。经筛选的有核细胞、粒细胞、巨噬细胞集落形成单位（CFU-GM）、红系爆发形成单位（BFU-E）以及粒细胞、红细胞、单核细胞和巨核细胞集落形成单位的平均回收率（+/-1 SD）使用与预冷冻样品相同的培养条件，CFU-GEMM分别为83 +/- 12、95 +/- 16、84 +/- 25和85 +/- 25。CFU-GM、BFU-E和CFU-GEMM的增殖能力是完整的，因为产生的集落分别含有高达22，500、182，500和292，500个细胞。CFU-GEMM的自我更新能力也得以保持，单个CFU-GEMM集落在20度培养皿中的重新接种率>96%，并获得CFU-GM、BFU-E和CFU-GEMM的20度集落。此外，解冻后通过FACS分离的CD 34（+）CD 38（-）细胞产生>250倍的HPC离体扩增。为了评估HSC的能力，将来自单个收集物的解冻物珠分离成CD 34（+）细胞，并输注到亚致死剂量照射的非肥胖糖尿病（NOD）/严重联合免疫缺陷（SCID）小鼠中。11-13周后，在小鼠中检测到具有多谱系表型的CD 45（+）人细胞植入;植入水平与新鲜CB报告的水平相当。因此，具有高增殖性、再铺板、离体扩增和小鼠NOD/SCID移植能力的未成熟人CB细胞可以冷冻储存> 15年，可以有效地回收，并且很可能对临床移植保持有效。
Transplanted cord blood (CB) hematopoietic stem cells (HSC) and progenitor cells (HPC) can treat malignant and nonmalignant disorders. Because long-term cryopreservation is critical for C8 banking and transplantation, we assessed the efficiency of recovery of viable HSC/HPC from individual CBs stored frozen for 15 yr. Average recoveries (+/-1 SD) of defrosted nucleated cells, colony-forming unit-granulocyte, -macrophage (CFU-GM), burst-forming unit-erythroid (BFU-E), and colony-forming unit-granulocyte, -erythrocyte, -monocyte, and -megakaryocyte (CFU-GEMM) were, respectively, 83 +/- 12, 95 +/- 16, 84 +/- 25, and 85 +/- 25 using the same culture conditions as for prefreeze samples. Proliferative capacities of CFU-GM, BFU-E, and CFU-GEMM were intact as colonies generated respectively contained up to 22,500,182,500, and 292,500 cells. Self-renewal of CFU-GEMM was also retained as replating efficiency of single CFU-GEMM colonies into 2degrees dishes was >96% and yielded 2degrees colonies of CFU-GM, BFU-E, and CFU-GEMM. Moreover, CD34(+)CD38(-) cells isolated by FACS after thawing yielded >250-fold ex vivo expansion of HPC. To assess HSC capability, defrosts from single collections were bead-separated into CD34(+) cells and infused into sublethally irradiated nonobese diabetic (NOD)/severe combined immunodeficient (SCID) mice. CD45(+) human cell engraftment with multilineage phenotypes was detected in mice after 11-13 wk; engrafting levels were comparable to that reported with fresh CB. Thus, immature human CB cells with high proliferative, replating, ex vivo expansion and mouse NOD/SCID engrafting ability can be stored frozen for > 15 yr, can be efficiently retrieved, and most likely remain effective for clinical transplantation.