Huang Yaojin, Xie Xiaowei, Liu Mengyao, Zhang Yawen, Yang Junye, Yang Wenling, Hu Yu, Qi Saibing, Feng Yahui, Liu Guojun, Lu Shihong, Peng Xuemei, Ye Jinhui, Ma Shihui, Sun Jiali, Wang Lu, Hu Linping, Wang Lin, Zhu Xiaofan, Cheng Hui, Sun Zimin, Chen Junren, Dong Fang, Zhang Yingchi, Cheng Tao
State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of the Cell Ecosystem, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China.
Tianjin Institutes of Health Science, Tianjin, China.
J Clin Invest. 2025 Mar 4;135(9). doi: 10.1172/JCI183607. eCollection 2025 May 1.
Umbilical cord blood (UCB) plays substantial roles in hematopoietic stem cell (HSC) transplantation and regenerative medicine. UCB is usually cryopreserved for years before use. It remains unclear whether and how cryopreservation affects UCB function. We constructed a single-cell transcriptomics profile of CD34+ hematopoietic stem and progenitor cells (HSPCs) and mononuclear cells (MNCs) from fresh and cryopreserved UCB stored for 1, 5, 10, and 19 years. Compared with fresh UCB, cryopreserved HSCs and multipotent progenitors (MPPs) exhibited more active cell-cycle and lower expression levels of HSC and multipotent progenitor signature genes. Hematopoietic reconstitution of cryopreserved HSPCs gradually decreased during the first 5 years but stabilized thereafter, aligning with the negative correlation between clinical neutrophil engraftment and cryopreservation duration of UCB. Cryopreserved HSPCs also showed reduced megakaryocyte generation. In contrast, cryopreserved NK cells and T cells maintained a capacity for cytokine production and cytotoxicity comparable to that of fresh cells. Mechanistically, cryopreserved HSPCs exhibited elevated ROS, reduced ATP synthesis, and abnormal mitochondrial distribution, which collectively led to attenuated hematopoietic reconstitution. These effects could be ameliorated by sulforaphane (SF). Together, we elucidate the negative effect of cryopreservation on UCB HSPCs and identify SF as a mitigation strategy, broadening the temporal window and scope for clinical applications of cryopreserved UCB.
脐带血（UCB）在造血干细胞（HSC）移植和再生医学中发挥着重要作用。UCB通常在使用前冷冻保存数年。目前尚不清楚冷冻保存是否以及如何影响UCB的功能。我们构建了来自新鲜和冷冻保存1年、5年、10年和19年的UCB的CD34+造血干细胞和祖细胞（HSPCs）及单核细胞（MNCs）的单细胞转录组图谱。与新鲜UCB相比，冷冻保存的HSCs和多能祖细胞（MPPs）表现出更活跃的细胞周期以及更低的HSC和多能祖细胞特征基因表达水平。冷冻保存的HSPCs的造血重建在前5年逐渐下降，但此后趋于稳定，这与临床中性粒细胞植入与UCB冷冻保存时间的负相关一致。冷冻保存的HSPCs还显示巨核细胞生成减少。相比之下，冷冻保存的NK细胞和T细胞维持了与新鲜细胞相当的细胞因子产生和细胞毒性能力。机制上，冷冻保存的HSPCs表现出活性氧升高、ATP合成减少以及线粒体分布异常，这些共同导致造血重建减弱。萝卜硫素（SF）可改善这些影响。总之，我们阐明了冷冻保存对UCB HSPCs的负面影响，并确定SF作为一种缓解策略，拓宽了冷冻保存UCB临床应用的时间窗口和范围。
