切换至 "中华医学电子期刊资源库"

中华细胞与干细胞杂志(电子版) ›› 2021, Vol. 11 ›› Issue (04) : 200 -206. doi: 10.3877/cma.j.issn.2095-1221.2021.04.002

论著

冻存前后人脐带间充质干细胞对T和B淋巴细胞免疫抑制能力的差异比较
曾伟杰1, 廖延2, 胡樾2, 胡隽源2, 曾桂芳2, 傅泽钦2, 伍世铎2, 梁晓3, 谢长峰1, 刘沐芸3,()   
  1. 1. 518000 深圳,个体化细胞治疗技术国家地方联合工程实验室 (深圳)
    2. 518000 深圳,深圳市北科生物科技有限公司
    3. 518000 深圳,个体化细胞治疗技术国家地方联合工程实验室 (深圳);518000 深圳,深圳市北科生物科技有限公司
  • 收稿日期:2020-07-14 出版日期:2021-08-01
  • 通信作者: 刘沐芸

Comparison of immunosuppressive ability of human umbilical cord mesenchymal stem cells against T and B lymphocytes before and after cryopreservation

Weijie Zeng1, Yan Liao2, Yue Hu2, Juanyuan Hu2, Guifang Zeng2, Zeqin Fu2, Shiduo Wu2, Xiao Liang3, Changfeng Xie1, Muyun Liu3,()   

  1. 1. Department of National-local Associated Engineering Laboratory for Personalized Cellular Therapy, Shenzhen 518000, China
    2. Shenzhen Beike Biotechnology Company Limited, Shenzhen 518000, China
    3. Department of National-local Associated Engineering Laboratory for Personalized Cellular Therapy, Shenzhen 518000, China; Shenzhen Beike Biotechnology Company Limited, Shenzhen 518000, China
  • Received:2020-07-14 Published:2021-08-01
  • Corresponding author: Muyun Liu
引用本文:

曾伟杰, 廖延, 胡樾, 胡隽源, 曾桂芳, 傅泽钦, 伍世铎, 梁晓, 谢长峰, 刘沐芸. 冻存前后人脐带间充质干细胞对T和B淋巴细胞免疫抑制能力的差异比较[J/OL]. 中华细胞与干细胞杂志(电子版), 2021, 11(04): 200-206.

Weijie Zeng, Yan Liao, Yue Hu, Juanyuan Hu, Guifang Zeng, Zeqin Fu, Shiduo Wu, Xiao Liang, Changfeng Xie, Muyun Liu. Comparison of immunosuppressive ability of human umbilical cord mesenchymal stem cells against T and B lymphocytes before and after cryopreservation[J/OL]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2021, 11(04): 200-206.

目的

探讨冻存前和复苏后人脐带间充质干细胞(hUCMSCs)对T和B淋巴细胞免疫抑制能力的差异。

方法

分离健康人外周血单个核细胞,使用Anti-CD3和Anti-CD28单克隆抗体体外激活T淋巴细胞,使用ODN2395,hCD40L,羊抗人IgM抗体和白介素2体外刺激经过分选的B淋巴细胞,通过冻存前后hUCMSCs进行干预处理,与活化后的T和B淋巴细胞进行共培养,分为单独刺激组(激活后的T或B淋巴细胞单独培养)与hUCMSCs共培养组(激活后的T或B淋巴细胞分别与hUCMSCs共培养),分析hUCMSCs对激活的T和B淋巴细胞的免疫抑制作用,探究冻存前后hUCMSCs免疫调节作用的差异。两组间比较采用独立样本t检验,多组间比较采用重复测量设计方差分析。

结果

与冻存前hUCMSCs对比,冻存后hUCMSCs在抑制CD3+T细胞(37.60﹪±0.54﹪比39.40﹪±1.57﹪)、CD4+T细胞(36.87﹪±0.54﹪比38.63﹪±1.39﹪)和CD8+T细胞增殖能力(40.37﹪±1.14﹪比42.47﹪±1.90﹪)比较,差异无统计学意义(P均> 0.05);在促进CD3+T (18.07﹪±0.66﹪比16.77﹪±1.15﹪)、CD4+T(26.47﹪±1.13﹪比24.60﹪±1.47﹪)和CD8+T细胞凋亡能力(3.52﹪±0.22﹪比2.72﹪ ± 0.06﹪),差异无统计学意义(P均> 0.05);增加Treg细胞群比例能力(5.51﹪±0.71﹪比6.87﹪±0.27﹪),差异无统计学意义(P > 0.05);降低Th1亚群(0.47﹪±0.09﹪比0.33﹪± 0.04﹪)和Th17亚群比例能力(0.21﹪±0.04﹪比0.22﹪±0.03﹪),差异无统计学意义(P均> 0.05);在对B淋巴细胞增殖能力(38.83﹪±0.25﹪比34.23﹪±4.12﹪),促进B淋巴细胞的凋亡能力(54.95﹪±1.45﹪比55.10﹪±3.80﹪),减少IgA [(1045.09± 99.24)ng/mL比(1014.31±102.55)ng/mL]和IgG抗体的分泌能力[(120.51±4.94) ng/mL比(127.77±7.94) ng/mL],差异无统计学意义(P均> 0.05)。而在降低Th2细胞亚群比例(0.73﹪ ± 0.07﹪比0.49﹪±0.06﹪)和降低IgM抗体分泌[(739.70±28.39)ng/mL比(560.21±3.81)ng/mL]的能力上,冻存前hUCMSCs的作用能力弱于复苏后hUCMSCs,差异具有统计学意义(P均< 0.05)。

结论

冻存后hUCMSCs在降低Th2细胞亚群比例和降低IgM抗体分泌的能力上强于冻存前hUCMSCs,但整体的免疫调节能力相当,因此冻存前后的hUCMSCs均具有很高的临床应用价值。

Objective

To explore the immunosuppressive ability of human umbilical cord mesenchymal stem cells (hUCMSCs) on T and B lymphocytes before and after cryopreservation and provide future for using the clinical application of hUCMSCs.

Methods

Human peripheral blood mononuclear cells were isolated from healthy human peripheral blood. Anti-CD3 and anti-CD8 monoclonal antibodies were used to activate T lymphocytes in vitro. Selected B lymphocytes were stimulated by ODN2395, human CD40 ligand, goat anti-human IgM antibodies and interleukin 2 in vitro. The cells were co-cultured with activated T and B lymphocytes, divided into a single stimulation group and hUCMSCs co-culture group (before and after cryopreservation) . The immunosuppressive ability of hUCMSCs on activated T and B lymphocytes were analyzed. In addition, the differences immunosuppressive ability of hUCMSCs before and after cryopreservation were also studied. The differences between groups were compared by t-test, and the differences among groups were compared by repeated measures design analysis of variance.

Results

Compared with hUCMSCs before cypropreservation, the proliferation of CD3+T cells (37.60﹪±0.54﹪vs 39.40﹪±1.57﹪) , CD4+T cells (36.87﹪±0.54﹪vs 38.63﹪±1.39﹪) and CD8+T cells (40.37﹪±1.14﹪vs 42.47﹪±1.90﹪) (P > 0.05) were not significantly inhibited after cryopreservation. The apoptosis of CD3+T cells (18.07﹪±0.66﹪vs 16.77﹪±1.15﹪) , CD4+T cells (26.47﹪±1.13﹪vs 24.60﹪±1.47﹪) and CD8+T cells (3.52﹪±0.22﹪vs 2.72﹪±0.06﹪) had no significant difference (P > 0.05) . The proportion of regulatory cells (Treg) subsets (5.51﹪±0.71﹪vs 6.87﹪± 0.27﹪) had also no significant difference (P > 0.05) . hUCMSCs did not reduce the proportion of helper T cell (Th) subsets, especially the proportion of Th1 subsets (0.47﹪± 0.09﹪vs 0.33﹪± 0.04﹪) and Th17 subsets (0.21﹪± 0.04﹪vs 0.22﹪± 0.03﹪) (P > 0.05) . For B lymphocytes, hUCMSCs reduced the proportion of Th2 cell subsets (0.73﹪±0.07﹪vs 0.49﹪±0.06﹪) and IgM antibody secretion [ (739.70±28.39) ng/mL vs (560.21±3.81) ng/mL] after cryopreservation (P < 0.05) . There was no significant difference in proliferation and apoptosis of B lymphocytes, and reduced secretion of IgA and IgG antibodies before and after hUCMSCs cytopreservation (P > 0.05) .

Conclusion

After cryopreservation, hUCMSCs were more powerful than those before cryopreservation in reducing the proportion of Th2 cell subsets and the secretion of IgM antibodies. However, the immunosuppressive abilities was same. Both hUCMSCs before and after cryopreservation have high clinical application value.

表1 冻存前后P3代hUCMSCs与PBMC共培养对T细胞亚群增殖的影响( ± s,﹪)
表2 冻存前后P3代hUCMSCs与PBMC共培养对T细胞亚群凋亡的影响( ± s,﹪)
表3 冻存前后P3代hUCMSCs与PBMC共培养后对Th1、Th2和Th17亚群的影响( ± s,﹪)
表4 冻存前后P3代hUCMSCs与B淋巴细胞共培养后对B细胞分泌Ig抗体的影响( ± s,ng/mL)
1
单佳柔, 尼贝贝, 李翠平, 等. 人脐带间充质干细胞通过线粒体转移减轻肝细胞缺血-再灌注损伤的机制研究[J]. 器官移植, 2021, 12(3):294-301.
2
Riordan NH, Morales I, Fernandez G, et al. Clinical feasibility of umbilical cord tissue-derived mesenchymal stem cells inthe treatment of multiple sclerosis[J]. J Transl Med, 2018, 16(1):57.
3
Bartolucci J, Verdugo FJ, Gonzalez PL, et al. Safety and efficacy of the intravenous infusion of umbilical cord mesenchymal stem cells in patients with heart failure: a phase 1/2 randomized controlled trial (RIMECARD Trial [Randomized Clinical Trial of Intravenous Infusion Umbilical Cord Mesenchymal Stem Cells on Cardiopathy])[J]. Circ Res, 2017, 121(10):1192-1204.
4
Matas J, Orrego M, Amenabar D, et al. Umbilical cord-derived mesenchymal stromal cells (MSCs) for knee osteoarthritis: repeated MSC dosing is superior to a single MSC dose and to hyaluronic acid in a controlled randomized phase I/II trial[J]. Stem Cells Transl Med, 2019, 8(3):215-224.
5
Le Blanc K, Tammik L, Sundberg B, et al. Mesenchymal stem cells inhibit and stimulate mixed lymphocyte cultures andmitogenic responses independently of the major histocompatibility complex[J]. Scand J Immunol, 2003, 57(1):11-20.
6
Naji A, Eitoku M, Favier B, et al. Biological functions of mesenchymal stem cells and clinical implications[J]. Cell Mol Life Sci, 2019, 76(17):3323-3348.
7
Zhou T, Li H, Liao C, et al. Clinical efficacy and safety of mesenchymal stem cells for systemic lupus erythematosus[J]. Stem Cells Int, 2020, 2020:6518508. doi: 10.1155/2020/6518508.
8
Wang L, Huang S, Li S, et al. Efficacy and safety of umbilical cord mesenchymal stem cell therapy for rheumatoid arthritis patients: a prospective phase I/II study[J]. Drug Des Devel Ther, 2019, 13:4331-4340.
9
Dulugiac M, Moldovan L, Zarnescu O. Comparative studies of mesenchymal stem cells derived from different cord tissue compartments-the influence of cryopreservation and growth media[J]. Placenta, 2015, 36(10):1192-1203.
10
Chen Y, Yu Q, Hu Y, et al. Current research and use of mesenchymal stem cells in the therapy of autoimmune diseases[J]. Curr Stem Cell Res Ther, 2019, 14(7):579-582.
11
Boberg E, von Bahr L, Afram G, et al. Treatment of chronic GvHD with mesenchymal stromal cells induces durable responses: a phase II study[J]. Stem Cells Transl Med, 2020, 9(10):1190-1202.
12
Matsushita K. Mesenchymal stem cells and metabolic syndrome: current understanding and potential clinical implications[J]. Stem Cells Int, 2016, 2016:2892840. doi: 10.1155/2016/2892840.
13
Atluri S, Manchikanti L, Hirsch JA. Expanded umbilical cord mesenchymal stem cells (UC-MSCs) as a therapeutic strategy in managing critically Ill COVID-19 patients: the case for compassionate use[J]. Pain physician, 2020, 23(2):E71-E83.
14
Luetzkendorf J, Nerger K, Hering J, et al. Cryopreservation does not alter main characteristics of good manufacturing process-grade human multipotent mesenchymal stromal cells including immunomodulating potential and lack of malignant transformation[J]. Cytotherapy, 2015, 17(2):186-198.
15
Moll G, Alm J J, Davies L C, et al. Do cryopreserved mesenchymal stromal cells display impaired immunomodulatory and therapeutic properties?[J]. Stem cells, 2014, 32(9):2430-2442.
16
Chinnadurai R, Copland IB, Garcia MA, et al. Cryopreserved mesenchymal stromal cells are susceptible to T-cell mediated apoptosis which is partly rescued by IFNγ licensing[J]. Stem cells, 2016, 34(9):2429-2442.
17
Comoli P, Ginevri F, Maccario R, et al. Human mesenchymal stem cells inhibit antibody production induced in vitro by allostimulation[J]. Nephrol Dial Transplant, 2008, 23(4):1196-1202.
18
Corcione A, Benvenuto F, Ferretti E, et al. Human mesenchymal stem cells modulate B-cell functions[J]. Blood, 2006, 107(1):367-372.
[1] 赵俊杰, 王玺玉, 黄鹏飞, 张兆坤, 蒲彦川, 赵海燕. 辅助性T细胞17/调节性T细胞平衡在骨关节炎的作用[J/OL]. 中华关节外科杂志(电子版), 2024, 18(02): 209-214.
[2] 罗晨, 宗开灿, 李世颖, 傅应亚. 微小RNA-199a-3p调控CD4T细胞表达参与肺炎支原体肺炎患儿免疫反应研究[J/OL]. 中华妇幼临床医学杂志(电子版), 2023, 19(05): 569-574.
[3] 陈观梅, 左璇, 廖宝林. 慢性乙型肝炎新型免疫治疗研究进展[J/OL]. 中华实验和临床感染病杂志(电子版), 2024, 18(01): 7-10.
[4] 叶俊杰, 胡波涌. 人类免疫缺陷病毒感染者限期骨折内植物手术并发症的影响因素[J/OL]. 中华实验和临床感染病杂志(电子版), 2023, 17(06): 423-430.
[5] 陈天, 李歆, 刘开政, 邓永强. 口腔钛种植体成骨性能的研究进展[J/OL]. 中华口腔医学研究杂志(电子版), 2024, 18(03): 200-205.
[6] 赖全友, 高远, 汪建林, 屈士斌, 魏丹, 彭伟. 三维重建技术结合腹腔镜精准肝切除术对肝癌患者术后CD4+、CD8+及免疫球蛋白水平的影响[J/OL]. 中华普外科手术学杂志(电子版), 2024, 18(06): 651-654.
[7] 崔键, 戴庆. 基于肝囊型包虫病所致过敏反应模型研究Treg细胞数量比例与过敏反应的关系[J/OL]. 中华普外科手术学杂志(电子版), 2023, 17(04): 427-430.
[8] 张礼刚, 邹志辉, 许顺, 蔡可可, 胡永涛, 梁朝朝. 酒精对慢性非细菌性前列腺炎中T淋巴细胞变化的影响研究[J/OL]. 中华腔镜泌尿外科杂志(电子版), 2024, 18(01): 74-81.
[9] 符梅沙, 周玉华, 李慧, 薛春颜. 淋巴细胞免疫治疗对复发性流产患者外周血T淋巴细胞亚群分布与PD1/PD-L1表达的影响及意义[J/OL]. 中华临床医师杂志(电子版), 2023, 17(06): 726-730.
[10] 陆天, 孙道萍. 调节性B细胞在多发性骨髓瘤中的研究进展[J/OL]. 中华诊断学电子杂志, 2024, 12(02): 133-137.
[11] 吕泉龙, 史文杰, 孙文国. 免疫检查点抑制剂在治疗转移性去势抵抗性前列腺癌中的研究进展[J/OL]. 中华诊断学电子杂志, 2024, 12(01): 69-72.
[12] 计超, 向群. 乙酰胆碱受体对急性呼吸窘迫综合征小鼠T细胞亚群和炎症因子的影响[J/OL]. 中华诊断学电子杂志, 2024, 12(01): 50-56.
[13] 孟丽君, 宋芹, 邵莉, 李健. 系统性红斑狼疮合并肺动脉高压患者外周血T淋巴细胞亚群水平变化及临床意义[J/OL]. 中华诊断学电子杂志, 2024, 12(01): 38-43.
[14] 范茹, 刘宇清, 胡晓榕, 王轶奇, 张芬, 岑星, 卜玉洁, 陈俊伟. 系统性红斑狼疮患者长链非编码RNA表达变化及其与CD8+T细胞相关性研究[J/OL]. 中华诊断学电子杂志, 2023, 11(03): 184-189.
[15] 孙冠超, 万军, 石卉. IgG相关食物不耐受与肠道免疫微环境相关性的研究进展[J/OL]. 中华胃肠内镜电子杂志, 2024, 11(03): 200-203.
阅读次数
全文


摘要