一种新型冷冻保护剂冻存组织和细胞的效果  

Effect of a novel cryoprotectant in tissues and cells

作　　者：王清芳 张芬 昌广萍 李子晗 邢岚 彭浩 曾秀萍 钟桂强 陈辉 刘波 刘振宇 梁晓 Wang Qingfang;Zhang Fen;Chang Guangping;Li Zihan;Xing Lan;Peng Hao;Zeng Xiuping;Zhong Guiqiang;Chen Hui;Liu Bo;Liu Zhenyu;Liang Xiao(Shenzhen Beike Biotechnology Co.,Ltd.,Shenzhen 5180601,Guangdong Province,China)

机构地区：[1]深圳市北科生物科技有限公司,广东省深圳市518061

出　　处：《中国组织工程研究》2025年第36期7816-7826,共11页Chinese Journal of Tissue Engineering Research

摘　　要：背景:冷冻保存技术能够使组织/细胞于低温环境中长久贮存并维持活性与功能的完整性,这对细胞治疗、组织工程及生物样本库的构建有重大意义。冷冻保护剂常含二甲基亚砜和血清,为规避二甲基亚砜的毒副作用、血清成分的复杂性以及免疫反应等问题,部分成品冷冻保护剂虽已上市,但面临着成本高、应用受限等诸多难题,因此,迫切需要研发出一种成分明晰且能够解决上述问题的冷冻保护剂。目的:旨在评估一种新型冷冻保护剂对不同来源组织和细胞冻存效果的影响。方法:将新型冷冻保护剂作为实验组,市售及广泛使用的冷冻保护剂作为对照组,分别应用于脐带华通氏胶组织、脐带间充质干细胞、脐血/外周血单个核细胞、NK细胞及CIK细胞冻存,从冻存前和复苏后的细胞形态、数量、活率、表面标志物、分化潜能、细胞杀伤毒性等多方面进行对比分析,确认新型冷冻保护剂的冻存效果及潜在的应用价值。结果与结论:(1)采用新型冷冻保护剂能实现冻存脐带华通氏胶组织复苏后间充质干细胞形态正常,在细胞复苏回收率、表面标志物、分化潜能方面,实验组与对照组均无显著性差异;(2)实验组和对照组脐血/外周血单个核细胞冷冻复苏后的细胞数量和活率无显著性差异,且实验组和对照组NK细胞/CIK细胞冷冻复苏后的细胞数量和活率同样无显著性差异;(3)对于脐血/外周血单个核细胞衍生分化的NK细胞,实验组和对照组CD56^(+)CD16^(+)细胞亚群比例无显著差异,对于脐血/外周血单个核细胞衍生分化的CIK细胞,实验组和对照组CD3^(+)CD8^(+)和CD3^(+)CD56^(+)细胞亚群比例无显著差异;(4)在细胞杀伤毒性方面,当免疫细胞和黑色素瘤细胞系Mel624效靶比为20∶1时,无论是脐血还是外周血单个核细胞衍生分化的NK细胞/CIK细胞,实验组和对照组细胞杀瘤活性无显著差异。结果表明:新�BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton’s jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton’s jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and

关 键 词：冷冻保护剂 脐带华通氏胶组织 脐带间充质干细胞 脐血单个核细胞 外周血单个核细胞 NK细胞 CIK细胞 

分 类 号：R459.9[医药卫生—治疗学] R318.52[医药卫生—临床医学] Q813.1[生物学—生物工程]