| 高雅萌,赵珂,赵雄伟,吕志春,李思雨,武云强,孙慧颖,高慧英,向慎思,李长燕.胎肝及骨髓来源造血干/祖细胞电离辐射敏感性的比较研究[J].中华放射医学与防护杂志,2023,43(8):588-594 |
| 胎肝及骨髓来源造血干/祖细胞电离辐射敏感性的比较研究 |
| Comparative study of the radiosensitivity of hematopoietic stem/progenitor cells derived from fetal liver and bone marrow |
| 投稿时间：2023-04-27 |
| DOI：10.3760/cma.j.cn112271-20230427-00132 |
| 中文关键词: 骨髓造血干/祖细胞 胎肝造血干/祖细胞 线粒体功能 重建造血 电离辐射敏感性 |
| 英文关键词:Bone marrow-derived hematopoietic stem and progenitor cells Fetal liver-derived hematopoietic stem and progenitor cells Mitochondrial function Reconstitution capacity Radiosensitivity |
| 基金项目:国家重点研发计划（2022YFA1103502） |
| 作者 | 单位 | E-mail | | 高雅萌 | 安徽医科大学基础医学院, 合肥 230032 | | | 赵珂 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 赵雄伟 | 解放军总医院第五医学中心, 北京 100071 | | | 吕志春 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 李思雨 | 安徽医科大学基础医学院, 合肥 230032 | | | 武云强 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 孙慧颖 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 高慧英 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 向慎思 | 中国人民解放军军事科学院军事医学研究院辐射医学研究所, 北京 100850 | | | 李长燕 | 安徽医科大学基础医学院, 合肥 230032 | fmmli@163.com |
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| 中文摘要: |
| 目的 研究电离辐射对胎肝与骨髓来源造血干/祖细胞（hematopoietic stem and progenitor cells，HSPCs）的影响差异。方法 取14.5 d小鼠胎肝及8周龄小鼠骨髓，分选c-Kit+细胞体外接受60Co单次5、10 Gy照射，检测HSPCs中细胞凋亡、线粒体活性氧（ROS）水平、集落形成能力、DNA损伤情况。将12只CD45.1背景的C57BL/6J小鼠按随机数表法分成骨髓组（BM）和胎肝组（FL），60Co全身照射，第1次4.5 Gy照射，间隔30 min后再进行第2次5 Gy剂量照射，6 h后进行移植，12周后检测嵌合率、谱系分化以及细胞周期。分选胎肝及骨髓的c-Kit+细胞检测线粒体压力。结果 与骨髓HSPCs相比，照射后胎肝HSPCs的细胞凋亡比例显著升高（t=16.21、12.27，P<0.05），ROS水平显著增加（t=68.72、18.89，P<0.05）；集落形成能力显著降低，在5 Gy时胎肝HSPCs成集落能力显著降低，照射剂量为10 Gy时完全不能形成集落（t=12.41、15.67、9.46，P<0.05）；γ-H2AX免疫荧光染色结果显示，照射后胎肝HSPCs的DNA损伤更加严重，形成的Foci数量显著升高（t=2.27、2.03，P<0.05）。这些结果表明，胎肝HSPCs对辐射更加敏感。移植结果显示，移植胎肝HSPCs的嵌合率低于骨髓细胞（t=5.84，P<0.05）；在移植嵌合小鼠的骨髓与脾脏中，胎肝细胞组髓系细胞的比例高于骨髓细胞组，提示胎肝HSPCs体内重建能力低于骨髓HSPCs，并且更容易向髓系分化。细胞周期检测结果显示胎肝组S期比例显著高于骨髓组（t=2.89，P<0.05）。线粒体压力结果显示胎肝HSPCs基础呼吸能力（t=39.19，P<0.05）、质子泄漏（t=6.64，P<0.05）、三磷酸腺苷（ATP）产生（t=9.33，P<0.05）、耦联效率（t=7.10，P<0.05）均高于骨髓c-Kit+细胞；呼吸储备能力（t=5.53，P<0.05）低于骨髓细胞。结论 利用多种方法对照射后胎肝及骨髓来源的HSPCs进行检测，明确了电离辐射对两种不同HSPCs功能的影响，为深入探索放射对不同发育阶段造血干细胞的影响奠定基础。 |
| 英文摘要: |
| Objective To investigate the difference in the radiation sensitivity of hematopoietic stem and progenitor cells (HSPCs) derived from fetal liver and bone marrow.Methods HSPCs from fetal liver of 14.5 d embryo or bone marrow of 8 week-old mice were isolated to receive a single dose of 5 or 10 Gy irradiation in vitro using a 60Co irradiator. Twelve hours later, the cell apoptosis, mitochondrial reactive oxygen species (ROS) level, colony formation ability and DNA damage in HSPCs were detected. Freshly isolated HSPCs were injected into lethally irradiated CD45.1+ C57BL/6J mice (4.5 Gy+5 Gy with an interval of 30 min) Chimerism rate, lineage constitution, and cell cycle were analyzed 12 weeks after transplantation.Results Compared with bone marrow HSPCs after irradiation, the percentage of apoptosis in fetal liver HSPCs was significantly higher (t=16.21, 12.27, P<0.05), the level of ROS was dramatically elevated (t=68.72, 18.89, P<0.05). At 10 Gy, fetal liver HSPCs could not form colonies at all (t=12.41, 15.67, 9.46, P<0.05). γ-H2AX immunofluorescence staining showed that the DNA damage of fetal liver HSPCs was more severe after irradiation, and the number of Foci formed was significantly higher than that of bone marrow HSPCs (t=2.27, 2.03, P<0.05), which indicated that fetal liver HSPCs were more sensitive to radiation. The chimerism rate of transplanted fetal liver HSPCs was lower than that of bone marrow cells (t=5.84, P<0.05) with a higher proportion of myeloid lineage, suggesting that fetal liver HSPCs had lower in vivo reconstitution capacity than bone marrow HSPCs and were more prone to myeloid differentiation. The cell cycle of bone marrow HSPCs from transplanted chimeric mice was examined, and the proportion of S-phase was significantly higher in the fetal liver group than that in the bone marrow group (t=2.89, P<0.05). Mitochondrial stress results showed that fetal liver HSPCs had higher basal respiratory capacity (t=39.19, P<0.05), proton leakage (t=6.64, P<0.05), ATP production (t=9.33, P<0.05), and coupling efficiency (t=7.10, P<0.05) than bone marrow c-Kit+ cells, while respiratory reserve capacity (t=5.53, P< 0.05) was lower than that of bone marrow c-Kit+ cells.Conclusions HSPCs derived from fetal liver display higher radiosensitivty compared with bone marrow HSPCs, laying the foundation for an in-depth illustration of the effects of radiation on hematopoietic stem cells at different developmental stages. |
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