CXCL2在巨核细胞分化过程中的功能研究

doi:10.19586/j.2095-2341.2022.0217

生物技术进展 ›› 2023, Vol. 13 ›› Issue (2): 257-263.DOI: 10.19586/j.2095-2341.2022.0217

• 研究论文 • 上一篇

CXCL2在巨核细胞分化过程中的功能研究

王迪¹^,²(), 赵艳红¹^,², 梁一鹏¹^,², 宋昊泽¹^,², 石莉红¹^,²(), 佟静媛¹^,²()

^1.中国医学科学院血液病医院（中国医学科学院血液学研究所），实验血液学国家重点实验室，国家血液系统疾病临床医学研究中心，细胞生态海河实验室，天津 300020
^2.天津医学健康研究院，天津 301600

收稿日期:2022-12-30 接受日期:2023-02-07 出版日期:2023-03-25 发布日期:2023-04-07
通讯作者: 石莉红,佟静媛
作者简介:王迪 E-mail: wangdi@ihcams.ac.cn；
基金资助:
国家重点研发计划(2022YFA1103503);国家自然基金委项目(82100152);医科院医学与健康科技创新工程项目(2021-I2M-1-073);中国医学科学院中央级公益性科研院所基本科研业务费专项资金(2022-RC180-04);细胞生态海河实验室创新基金项目(22HHXBSS00017)

Research on the Function of CXCL2 During Megakaryocyte Differentiation

Di WANG¹^,²(), Yanhong ZHAO¹^,², Yipeng LIANG¹^,², Haoze SONG¹^,², Lihong SHI¹^,²(), Jingyuan TONG¹^,²()

^1.State Key Laboratory of Experimental Hematology，National Clinical Research Center for Blood Diseases，Haihe Laboratory of Cell Ecosystem，Institute of Hematology & Blood Diseases Hospital，Chinese Academy of Medical Sciences & Peking Union Medical College，Tianjin 300020，China
^2.Tianjin Institutes of Health Science，Tianjin 301600，China

Received:2022-12-30 Accepted:2023-02-07 Online:2023-03-25 Published:2023-04-07
Contact: Lihong SHI,Jingyuan TONG

摘要/Abstract

摘要：

临床中血小板输注供需矛盾愈发尖锐，体外血小板生成的相关研究在全球受到广泛关注，主要集中于干细胞来源、细胞因子、转录因子、湍流体系等方面。由于血小板由成熟巨核细胞释放产生，提高巨核细胞分化效率，将有利于体外生产血小板体系的优化。CXCL2具有促炎、促进血管生成的作用，与心血管疾病、结肠癌进展等有关，但其在巨核细胞分化过程中的功能尚不清楚。利用脐带血CD34⁺细胞向巨核细胞诱导分化体系，通过慢病毒介导的基因敲降方法降低CXCL2的表达水平，并利用流式细胞术检测巨核细胞的分化效率。结果表明，CXCL2被敲降后，CD34⁺细胞向巨核细胞的早期增殖分化受到抑制，并且这一抑制效应在增殖分化过程中持续存在。综上，CXCL2在巨核分化过程中发挥着重要的调控作用，这一结论将对体外巨核分化及生产血小板具有一定的指导意义。

关键词: 巨核细胞分化, CXCL2, CD34⁺细胞

Abstract:

Clinically， the conflict between supply and demand of platelet transfusion is becoming increasingly acute， and research related to platelet generation in vitro has received extensive attention globally， mainly focusing on stem cell sources， cytokines， transcription factors， and turbulent flow systems. As platelets are small fragments produced from megakaryocytes， promoting differentiation efficiency and production of megakaryocytes will optimize platelet generation in vitro. CXCL2 is a chemoattractant chemokine promoting inflammation and angiogenesis， which has been known to be associated with cardiovascular disease and colon cancer， but its function in megakaryocyte differentiation remains unclear. In this study， utilizing a model of differentiation from cord blood-derived CD34⁺ cells to megakaryocytes， CXCL2 expression was decreased by lentivirus-mediated knockdown method， and the differentiation efficiency of megakaryocytes was detected by flow cytometry. Results indicated that early stage of megakaryocyte differentiation was inhibited after CXCL2 knockdown， and this inhibitory effect persisted throughout the differentiation process. This study suggested that CXCL2 plays an important regulatory role in megakaryocyte differentiation， and this finding may exert some implications on megakaryocyte differentiation and platelet generation in vitro.

Key words: megakaryocyte differentiation, CXCL2, CD34⁺ cells

中图分类号:

Q254

王迪, 赵艳红, 梁一鹏, 宋昊泽, 石莉红, 佟静媛. CXCL2在巨核细胞分化过程中的功能研究[J]. 生物技术进展, 2023, 13(2): 257-263.

Di WANG, Yanhong ZHAO, Yipeng LIANG, Haoze SONG, Lihong SHI, Jingyuan TONG. Research on the Function of CXCL2 During Megakaryocyte Differentiation[J]. Current Biotechnology, 2023, 13(2): 257-263.

图/表 5

表1 本研究中的shRNA序列

Table 1 shRNA sequences in this study

引物名称	shRNA序列（5'→3')
Scrambled-F	5'-GTGCGTTGTTAGTACTAATCCTATTT-3'
Scrambled-R	5'-AAATAGGATTAGTACTAACAACGCAC-3'
CXCL2-sh1F	5'-GCATCGCCCATGGTTAAGAAA-3'
CXCL2-sh1R	5'-TTTCTTAACCATGGGCGATGC-3'
CXCL2-sh2F	5'-ACACTCAAGAATGGGCAGAAA-3'
CXCL2-sh2R	5'-TTTCTGCCCATTCTTGAGTGT-3'
CXCL2-sh3F	5'-GCAGATATTCTCTAGTCATTT-3'
CXCL2-sh3R	5'-AAATGACTAGAGAATATCTGC-3'
CXCL2-sh4F	5'-ATTTCTTCGTGATGACATATC-3'
CXCL2-sh4R	5'-GATATGTCATCACGAAGAAAT-3'

图1 脐带血CD34+细胞向巨核细胞诱导分化实验模式图

Fig. 1 Schematic illustration of megakaryocyte differentiation of cord blood

图2 CXCL2基因在人类巨核细胞分化各个阶段的表达情况注：脐带血来源的血细胞RNA Microarray数据。HSC—造血干细胞；CMP—常见髓系祖细胞；MEP—巨核细胞-红细胞祖细胞；CFU-MK—巨核细胞集落形成单位；MK—巨核细胞。

Fig. 2 The expression of CXCL2 in each stage of megakaryocyte differentiation

图3 HEL细胞及脐带血CD34+细胞中CXCL2的检测A：通过RT-qPCR检测HEL细胞中CXCL2的敲降效率；B：通过RT-qPCR检测脐带血CD34+细胞中CXCL2的敲降效率

Fig. 3 Detection of CXCL2 knockdown in HEL cells and cord blood CD34+ cells

图4 敲降CXCL2基因对脐带血CD34+细胞向巨核细胞诱导分化的影响A：流式细胞术检测脐带血CD34+细胞向巨核细胞诱导分化5、8、11 d后各个细胞亚群的比例；B～D：统计学分析脐带血CD34+细胞向巨核诱导分化5、8、11 ｄ后CD41a+细胞、CD41a+CD42b-细胞、CD41a+CD42b+细胞比例；E：11 ｄ时CD41a+CD42b+细胞中多倍体（≥4N）细胞的比例；F：11 ｄ时显微镜下测量的CD41a+CD42b+细胞直径大小；G：11 ｄ时瑞氏-姬姆萨染色分析CD41a+CD42b+细胞形态（比例尺为10 μm）。*表示在P<0.05水平上具有统计学意义，**表示在P<0.01水平上具有统计学意义，***表示在P<0.001水平上具有统计学意义，****表示在P<0.000 1水平上具有统计学意义，ns表示P>0.05水平上无统计学差异

Fig. 4 The effect of CXCL2 knockdown on megakaryocyte differentiation from cord blood CD34+ cells

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CXCL2在巨核细胞分化过程中的功能研究

Research on the Function of CXCL2 During Megakaryocyte Differentiation

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