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中华细胞与干细胞杂志(电子版)

中华细胞与干细胞杂志(电子版) ›› 2020, Vol. 10 ›› Issue (02) : 82 -89. doi: 10.3877/cma.j.issn.2095-1221.2020.02.003

所属专题： 文献；

论著

UM171和SR1对不同来源造血干/祖细胞体外扩增的作用研究
董忱1, 文瑞婷1, 杨扬2, 赵龙2, 王丽3, 段炼4, 陈虎2, 张斌2,†()   
  1. 1. 100850　北京，军事科学院军事医学研究院；100071　北京，解放军总医院第五医学中心造血干细胞移植科；100071　北京，造血干细胞治疗及转化研究北京市重点实验室
    2. 100071　北京，解放军总医院第五医学中心造血干细胞移植科；100071　北京，造血干细胞治疗及转化研究北京市重点实验室
    3. 100071　北京，解放军总医院妇产科
    4. 100071　北京，解放军总医院神经外科
  • 收稿日期:2019-12-13 出版日期:2020-04-01
  • 通信作者: 张斌
  • 基金资助:
    北京市科技计划课题(Z181100001818004)

Effects of UM171 and SR1 on ex vivo expansion of hematopoietic stem and progenitor cells from different sources

Chen Dong1, Ruiting Wen1, Yang Yang2, Long Zhao2, Li Wang3, Lian Duan4, Hu Chen2, Bin Zhang2,†()   

  1. 1. Academy of Military Medical Sciences, Academy of Military Science, Beijing 100850, China; Department of Hematopoietic Stem Cell Transplantation, the Fifth Medical Center of the PLA General Hospital, Beijing 100071, China; Beijing Key Laboratory of Stem Cell Therapy and Transformation Research, Beijing 100071, China
    2. Department of Hematopoietic Stem Cell Transplantation, the Fifth Medical Center of the PLA General Hospital, Beijing 100071, China; Beijing Key Laboratory of Stem Cell Therapy and Transformation Research, Beijing 100071, China
    3. Department of Obstetrics and Gynecology, the Fifth Medical Center of the PLA General Hospital, Beijing 100071, China
    4. Department of Neurosurgery, the Fifth Medical Center of the PLA General Hospital, Beijing 100071, China
  • Received:2019-12-13 Published:2020-04-01
  • Corresponding author: Bin Zhang
  • About author:
    Corresponging author:Zhang Bin, Email:
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引用本文：

董忱, 文瑞婷, 杨扬, 赵龙, 王丽, 段炼, 陈虎, 张斌. UM171和SR1对不同来源造血干/祖细胞体外扩增的作用研究[J]. 中华细胞与干细胞杂志(电子版), 2020, 10(02): 82-89.

Chen Dong, Ruiting Wen, Yang Yang, Long Zhao, Li Wang, Lian Duan, Hu Chen, Bin Zhang. Effects of UM171 and SR1 on ex vivo expansion of hematopoietic stem and progenitor cells from different sources[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2020, 10(02): 82-89.

目的

探讨小分子化合物UM171和SR1对脐带血、供者动员外周血和淋巴瘤患者自体动员外周血3种来源的造血干/祖细胞（HSPCs）体外扩增的作用。

方法

将3种来源的CD34+细胞分别予以UM171、SR1干预后进行体外扩增培养，记为对照组、UM171组、SR1组和UM171+SR1组。通过细胞计数检测各组总有核细胞的数量，流式细胞术检测HSPCs的比例、各谱系分化细胞的比例和HSPCs上归巢相关因子CXCR4的表达水平。多组数据若满足方差齐性，采用单因素方差分析，组间两两比较采用LSD-t检验；若方差不齐，多组间比较以及两两比较均采用Kruskal-Wallis检验。

结果

与对照组比较，UM171和SR1均能促进3种来源HSPCs的比例升高，同时UM171能够增加3种来源HSPCs的扩增倍数。与对照组比较，UM171处理后脐带血来源的CD33+ （髓系）细胞的比例升高，CD41+ （巨核）细胞的比例降低；SR1处理后3种来源的CD3-CD56+ （自然杀伤）细胞的比例均升高。体外扩增48 h后各组HSPCs上CXCR4的表达较培养前增加。

结论

UM171能够有效扩增3种来源HSPCs的数量，促进脐带血来源HSPCs分化为髓系细胞并抑制其分化为巨核细胞。SR1能够促进3种来源HSPCs分化为自然杀伤细胞。体外扩增培养可以提高3种来源HSPCs上CXCR4的表达水平。

关键词: 小分子化合物, 造血干/祖细胞, 扩增, 分化, CXCR4
Objective

To investigate the effects of small molecule compounds UM171 and SR1 on hematopoietic stem and progenitor cells (HSPCs) from umbilical cord blood (CB) , donor mobilized peripheral blood (Donor-mPB) and autologous mobilized peripheral blood (Auto-mPB) .

Methods

HSPCs from three sources were cultured with or without UM171 and SR1, and named control group, UM171 group, SR1 group and UM171+SR1 group.The number of total nucleated cells in each group was detected by cell counting. The proportion and differentiation of HSPCs and the expression of CXCR4 on HSPCs were detected by flow cytometry.One-way ANOVA and Kruskal- Wallis test were used for statistical analysis.

Results

Compared with control group, the proportions of HSPCs from three sources were increased both after treatment of UM171 and SR1. And the treatment of UM171 could increase the expansion folds of HSPCs from three sources significantly. Compared with control group, UM171 treatment led an increase in the proportion of CD33+ (myeloid) cells as well as a decrease in the proportion of CD41+ (megakaryocyte) cells from CB-derived HSPCs, SR1 treatment led an increase in the proportion of CD3- CD56+ (natural killer) cells from HSPCs from three sources. The expression of CXCR4 on HSPCs in each group increased significantly after 48 h of ex vivo expansion relative to that before culture.

Conclusion

UM171 could effectively expand the number of HSPCs from CB, Donor-mPB, Auto-mPB and promote the differentiation of CB-derived HSPCs into myeloid cells as well as inhibit their differentiation into megakaryocytes. SR1 could promote the differentiation of HSPCs from three sources into natural killer cells. The expression of CXCR4 on HSPCs from three sources was increased significantly after ex vivo expansion.

Key words: Small molecule compounds, Hematopoietic stem and progenitor cells, Ex vivo expansion, Differentiation, CXCR4
表1 CB来源HSPCs体外扩增前后的比例（﹪， ± s，n = 5）
分组 CD34+细胞 CD34+ CD38-细胞 CD34+ CD38- CD45RA- CD90+细胞
未培养组 93.64±1.63 10.19±2.48 1.82±1.61
对照组 52.57±5.98a 39.31±4.62a 0.80±0.54
UM171组 80.12±4.83ab 68.02±7.72ab 3.71±3.06
SR1组 77.24±6.65ab 49.81±9.81abc 1.65±0.83
UM171+SR1组 87.91±4.20bcd 68.90±9.21abd 3.93±2.53
F值 50.291 54.634 2.423
P值 < 0.001 < 0.001 0.082
表2 CB来源HSPCs的扩增倍数（ ± s，n = 5）
分组 总有核细胞 CD34+细胞 CD34+ CD38-细胞 CD34+CD38-CD45RA- CD90+细胞
对照组 129.06±21.71 72.47±14.41 532.59±210.08 67.43± 30.84
UM171组 140.68±50.75 121.82±49.59 1037.36±587.09 318.53±147.94
SR1组 147.32±23.20 120.40±15.16 792.11±388.75 201.71±157.55
UM171+SR1组 167.54±69.82 156.97±65.79a 1249.83±670.19 473.88±356.18a
F/H值 1.344 8.714 5.000 3.420
P值 0.719 0.033 0.172 0.043
表4 Donor-mPB来源HSPCs的扩增倍数（ ± s，n = 5）
分组 总有核细胞 CD34+细胞 CD34+ CD38-细胞 CD34+ CD38- CD45RA- CD90+细胞
对照组 62.26±19.99 26.69± 9.28 216.19± 59.94 27.55±13.44
UM171组 69.57±25.32 58.38±27.42 543.13±242.33a 205.13±84.14a
SR1组 58.58±22.68 52.73±28.87 331.26±101.79 70.40±23.04b
UM171+SR1组 54.35±22.70 54.58±26.23 435.45±192.72 169.92±70.94ac
F/H值 0.402 1.773 8.166 10.779
P值 0.753 0.193 0.043 < 0.001
表6 Auto-mPB来源HSPCs的扩增倍数（ ± s，n = 5）
分组 总有核细胞 CD34+细胞 CD34+ CD38-细胞 CD34+ CD38-CD45RA- CD90+细胞
对照组 41.90±16.77 13.13± 5.83 204.62±129.92 19.98±13.89
UM171组 36.92±10.09 23.91± 6.54 448.34±136.22a 45.89±22.40
SR1组 33.58±13.24 21.00± 7.35 237.66±131.33b 37.72±27.34
UM171+SR1组 31.70±14.82 24.31±11.38 318.11±108.56 57.57±18.07
F值 0.514 2.067 3.650 2.822
P值 0.678 0.145 0.035 0.072
表3 Donor-mPB来源HSPCs体外扩增前后的比例（﹪， ± s，n = 5）
分组 CD34+细胞 CD34+ CD38-细胞 CD34+ CD38-CD45RA- CD90+细胞
未培养组 91.10±9.97 7.86±2.45 0.56±0.18
对照组 38.52±3.55a 26.94±4.10a 0.25±0.11
UM171组 74.02±5.09ab 56.86±6.99ab 1.55±0.29b
SR1组 77.47±2.94ab 44.08±5.21abc 0.68±0.24
UM171+SR1组 88.45±4.59bcd 60.33±6.01abd 1.76±0.67b
F/H值 66.041 88.635 20.012
P值 < 0.001 < 0.001 < 0.001
表5 Auto-mPB来源HSPCs体外扩增前后的比例（﹪， ± s，n = 5）
分组 CD34+细胞 CD34+ CD38-细胞 CD34+CD38-CD45RA- CD90+细胞
未培养组 98.18±2.92 3.70± 1.07 0.99±0.21
对照组 30.97±7.13a 17.39± 7.46a 0.51±0.35
UM171组 64.17±5.43ab 44.23± 8.36ab 1.28±0.69
SR1组 62.41±6.73ab 25.25± 9.03ac 1.25±0.78
UM171+SR1组 76.45±6.55abcd 39.73±10.45abd 2.16±1.20ab
F值 84.257 21.522 3.387
P值 < 0.001 < 0.001 0.029
表7 CB来源HSPCs体外扩增后各谱系分化细胞的比例（﹪， ± s，n = 5）
分组 CD33+细胞 CD41+细胞 CD235a+细胞 CD19+细胞 CD3+细胞 CD3-CD56+细胞
对照组 88.79±4.97 9.18±4.04 0.06±0.05 0.21±0.13 0.19±0.17 3.65±0.66
UM171组 97.73±1.53a 2.63±2.19a 0.08±0.07 0.13±0.12 0.18±0.13 3.69±0.83
SR1组 92.14±2.17b 4.71±1.99a 0.06±0.05 0.21±0.18 0.14±0.10 8.50±1.93ab
UM171+SR1组 96.15±1.16ac 3.02±1.04a 0.06±0.05 0.05±0.02 0.09±0.08 9.29±2.24ab
F/H值 9.813 6.882 0.183 7.746 0.796 18.736
P值 0.001 0.003 0.906 0.052 0.514 < 0.001
表8 Donor-mPB来源HSPCs体外扩增后各谱系分化细胞的比例（﹪， ± s，n = 5）
分组 CD33+细胞 CD41+细胞 CD235a+细胞 CD19+细胞 CD3+细胞 CD3-CD56+细胞
对照组 90.86±4.69 9.49±4.20 0.06±0.04 0.15±0.13 0.30±0.24 2.48±0.75
UM171组 95.86±2.00 12.91±5.77 0.08±0.05 0.10±0.07 0.19±0.15 2.38±1.08
SR1组 94.37±2.54 5.65±1.82 0.05±0.04 0.17±0.11 0.19±0.14 10.48±4.48ab
UM171+SR1组 94.35±1.68 8.64±3.22 0.03±0.02 0.13±0.10 0.08±0.07 10.79±3.54ab
F/H值 5.733 2.760 2.616 0.420 3.746 13.047
P值 0.125 0.076 0.455 0.741 0.290 < 0.001
表9 Auto-mPB来源HSPCs体外扩增后各谱系分化细胞的比例（﹪， ± s，n = 5）
分组 CD33+细胞 CD41+细胞 CD235a+细胞 CD19+细胞 CD3+细胞 CD3-CD56+细胞
对照组 94.34±1.51 6.12±1.16 0.12±0.11 0.15±0.07 0.15±0.12 3.03±2.19
UM171组 96.92±1.50 10.37±3.58 0.10±0.08 0.08±0.04 0.17±0.08 3.84±2.45
SR1组 95.70±3.37 3.54±1.06b 0.06±0.05 0.09±0.08 0.16±0.10 9.76±4.32ab
UM171+SR1组 95.98±1.67 4.38±0.44b 0.08±0.05 0.07±0.06 0.13±0.07 10.22±5.08ab
F值 1.217 15.434 0.567 1.450 0.142 5.234
P值 0.336 0.001 0.644 0.265 0.933 0.010
表10 CB来源HSPCs上CXCR4的表达水平（﹪， ± s，n = 3）
分组 CD34+CXCR4+细胞 CD34+ CD38-CXCR4+细胞 CD34+ CD38- CD90+CXCR4+细胞
未培养组 4.77±1.40 3.03±1.31 4.90±3.58
对照组 43.67±3.45a 39.27±1.86a 45.70±4.39a
UM171组 43.03±3.57a 37.90±2.75a 41.77±2.93a
SR1组 39.60±5.75a 37.53±3.30a 40.03±6.13a
UM171+SR1组 37.63±4.81a 35.13±4.18a 38.57±6.05a
F值 48.656 87.369 35.977
P值 < 0.001 < 0.001 < 0.001
表11 Donor-mPB来源HSPCs上CXCR4的表达水平（﹪， ± s，n = 3）
分组 CD34+CXCR4+细胞 CD34+ CD38-CXCR4+细胞 CD34+ CD38- CD90+CXCR4+细胞
未培养组 4.60±3.63 1.90±1.67 3.53±0.75
对照组 39.30±4.77a 40.10±5.00a 44.87±5.11a
UM171组 37.83±4.70a 48.77±5.95a 43.43±6.35a
SR1组 36.33±2.32a 40.03±3.35a 47.17±4.19a
UM171+SR1组 37.33±3.46a 40.33±2.57a 49.43±4.90a
F值 43.878 53.280 51.101
P值 < 0.001 < 0.001 < 0.001
表12 Auto-mPB来源HSPCs上CXCR4的表达水平（﹪， ± s，n = 3）
分组 CD34+CXCR4+细胞 CD34+ CD38-CXCR4+细胞 CD34+ CD38- CD90+CXCR4+细胞
未培养组 2.63± 1.75 1.23± 0.86 6.77±3.58
对照组 34.23± 4.72a 32.90± 6.30a 38.67±6.15a
UM171组 34.57± 6.17a 33.63± 7.80a 39.93±6.89a
SR1组 32.43±10.31a 32.33±13.63a 36.20±4.08a
UM171+SR1组 34.77±10.09a 35.23±13.63a 38.60±4.25a
F值 10.918 6.657 22.738
P值 0.001 0.007 < 0.001
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