生理和应激状态下骨髓与脾脏EBI的富集与分析

doi:10.19586/j.2095-2341.2026.0026

生物技术进展 ›› 2026, Vol. 16 ›› Issue (3): 686-699.DOI: 10.19586/j.2095-2341.2026.0026

生理和应激状态下骨髓与脾脏EBI的富集与分析

程启梅¹(), 王竟苇¹, 郑凌岳¹, 李鑫雨¹, 高程杰¹, 金煦¹, 高洁¹, 石莉红¹^,²(), 佟静媛¹()

^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），北京协和医学院，血液与健康全国重点实验室，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:2026-02-06 接受日期:2026-04-15 出版日期:2026-05-25 发布日期:2026-07-14
通讯作者: 石莉红,佟静媛
作者简介:程启梅 E-mail: chengqimei@ihcams.ac.cn
基金资助:
国家重点研发计划项目(2025YFA1804701);国家自然科学基金项目(82300141);国家自然科学基金项目(82500165);国家自然科学基金项目(82400148);天津市科技计划项目(24ZXRKSY00010);北京协和医学院中央高校基本科研业务费(3332025060);北京协和医学院中央高校基本科研业务费(3332025180);医科院医学与健康科技创新工程项目(2025-I2M-XHXX-136);医科院医学与健康科技创新工程项目(2025-I2M-XHXX-129)

Comparative Analysis of Erythroblastic Island in Bone Marrow Versus Spleen Under Steady-state and Stress Conditions

Qimei CHENG¹(), Jingwei WANG¹, Lingyue ZHENG¹, Xinyu LI¹, Chengjie GAO¹, Xu JIN¹, Jie GAO¹, Lihong SHI¹^,²(), Jingyuan TONG¹()

^1.State Key Laboratory of Experimental Hematology，National Clinical Research Center for Blood Diseases，Tianjin Key Laboratory of Cell Therapy 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:2026-02-06 Accepted:2026-04-15 Online:2026-05-25 Published:2026-07-14
Contact: Lihong SHI,Jingyuan TONG

摘要/Abstract

摘要：

为解析生理与应激状态下小鼠骨髓和脾脏中红系造血岛（erythroblastic island，EBI）的形态特征及细胞组成变化，并阐明髓外造血（extramedullary hematopoiesis，EMH）的微观结构基础。采用健康成年C57BL/6J小鼠作为实验对象，分别建立生理盐水对照组和苯肼（phenylhydrazine, PHZ）诱导急性溶血性贫血组，通过密度梯度沉降法富集骨髓与脾脏红系造血岛，并用流式细胞术分析细胞组成，成像流式观察形态及数量变化。结果显示，EBI红系造血岛核心组分包括红系前体细胞、巨噬细胞及粒/单核细胞，偶见淋巴细胞附着。生理与溶血状态下骨髓红系造血岛形态及各阶段红系细胞比例无显著差异。生理条件下成年小鼠脾脏仅含少量红系造血岛；急性溶血后脾脏体积增大2倍以上，红系造血岛数量显著增加，典型“花环样”结构密集出现，但岛内各阶段红系细胞构成比例稳定，原红细胞、早幼红细胞、中幼红细胞及网织红细胞分布模式与骨髓一致。这些结果提示脾脏通过红系造血岛数量扩增代偿性增加红细胞生成，岛内红系前体细胞仍遵循固有分化秩序，可为理解髓外造血的细胞学机制提供实验依据。

关键词: 红细胞生成, 红系造血岛, 脾脏, 骨髓, 应激造血

Abstract:

To investigate the morphology and cellular composition of erythroblastic island （EBI） in the bone marrow and spleen of mice under physiological and stress conditions， and to elucidate the microstructural basis of extramedullary hematopoiesis （EMH）. Healthy adult C57BL/6J mice were assigned to a saline-treated control group and a phenylhydrazine （PHZ）-induced acute hemolytic anemia group. EBI from the bone marrow and spleen were enriched via density gradient sedimentation. Their cellular composition was analyzed by flow cytometry， and their morphology and abundance were assessed using imaging flow cytometry. The results showed that EBI primarily consisted of erythroid precursor cells， macrophages， and granulocytic/monocytic cells， with occasional lymphocyte association. No significant differences were observed in bone marrow EBI morphology or in the proportions of erythroid cells at different stages between physiological and hemolytic conditions. Under physiological conditions， the spleen of adult mice contained only a small number of EBI； following acute hemolytic stress， spleen weight increased by more than twofold， the number of EBIs increased markedly， and densely distributed "rosette-like" structures were observed， yet the proportions of erythroid cells at different stages within the islands remained stable， with proerythroblasts， basophilic erythroblasts， polychromatic erythroblasts， and reticulocytes exhibiting distribution patterns consistent with those in the bone marrow. These findings demonstrated that the spleen compensates for increased erythrocyte production through quantitative expansion of EBI， while erythroid precursors within the islands maintain their intrinsic differentiation program， thereby providing experimental evidence for understanding the cellular mechanisms of EMH.

Key words: erythropoiesis, erythroblastic island, spleen, bone marrow, stress hematopoiesis

中图分类号:

Q291

程启梅, 王竟苇, 郑凌岳, 李鑫雨, 高程杰, 金煦, 高洁, 石莉红, 佟静媛. 生理和应激状态下骨髓与脾脏EBI的富集与分析[J]. 生物技术进展, 2026, 16(3): 686-699.

Qimei CHENG, Jingwei WANG, Lingyue ZHENG, Xinyu LI, Chengjie GAO, Xu JIN, Jie GAO, Lihong SHI, Jingyuan TONG. Comparative Analysis of Erythroblastic Island in Bone Marrow Versus Spleen Under Steady-state and Stress Conditions[J]. Current Biotechnology, 2026, 16(3): 686-699.

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链接本文: https://swjsjz.magtechjournal.com/CN/10.19586/j.2095-2341.2026.0026

https://swjsjz.magtechjournal.com/CN/Y2026/V16/I3/686

图/表 7

图1 显微镜观察瑞氏-吉姆萨染色的富集产物A：骨髓沉降前甩片图；B：骨髓沉降一次甩片图；C：骨髓沉降二次甩片图；D：脾脏沉降前甩片图；E：脾脏沉降一次甩片图；F：脾脏沉降二次甩片图

Fig. 1 Light microscopic analysis of Wright-Giemsa stained enriched products

图2 成年稳态造血时骨髓和脾脏中的EBI的细胞组成A：骨髓和脾脏中F4/80+的巨噬细胞在富集前后的细胞比例；B：骨髓和脾脏中F4/80+细胞的流式图；C：骨髓和脾脏中Ter119+的细胞在富集前后的细胞比例；D：骨髓和脾脏中Ter119+细胞的流式图；E：骨髓和脾脏中红系各阶段细胞在富集前的比例；F：骨髓和脾脏中红系各阶段细胞在富集前后的流式图；G：骨髓和脾脏中红系各阶段细胞在富集后的比例；H：骨髓和脾脏CD11b+ Gr-1+细胞在富集前后的比例；I：骨髓和脾脏CD11b+ Gr-1low细胞在富集前后的比例；J：骨髓和脾脏CD11b+ Gr-1hi细胞在富集前后的比例；K：骨髓和脾脏中粒/单核细胞在富集前后的流式图；L：骨髓和脾脏B220hi细胞在富集前后的比例；M：骨髓和脾脏B220low细胞在富集前后的比例；N：骨髓和脾脏中B细胞在富集前后的流式图；O：骨髓和脾脏T细胞在富集前后的比例；P：骨髓和脾脏中T细胞在富集前后的流式图。*、**、***表示差异在P0.05、P0.01、P0.001水平上具有统计学意义；ns表示无统计学意义。

Fig. 2 Cellular composition of EBI in bone marrow and spleen during steady-state hematopoiesis in adult mice

图3 成像流式分析骨髓与脾脏细胞团的结构差异及细胞组成特征A：正常骨髓样本圈取明场面积400的群体为R1；B：BM EBI中F4/80+的事件占R1的比例；C：BM EBI中CD71+的事件占R1的比例；D：BM EBI中F4/80+且CD71+的事件占R1的比例；E：BM EBI中CD11b+且CD71+的事件占R1的比例；F：BM EBI中CD11b+且F4/80+的事件占R1的比例；G：BM EBI中B220+且F4/80+的事件占R1的比例；H：BM EBI中各事件占R1的比例；I：BM EBI的影像；J：SP EBI的影像；K：正常脾脏样本圈取明场面积400的群体为R1；L：SP EBI中F4/80+的事件占R1的比例；M：SP EBI中CD71+的事件占R1的比例；N：SP EBI中F4/80+且CD71+的事件占R1的比例；O：SP EBI中CD11b+且CD71+的事件占R1的比例；P：SP EBI中CD11b+且F4/80+的事件占R1的比例；Q：SP EBI中B220+且F4/80+的事件占R1的比例；R：SP EBI中各事件占R1的比例。*、**、***表示差异在 P0.05、 P0.01、P0.001水平上具有统计学意义。

Fig. 3 Structural and compositional characterization of bone marrow and splenic cell clusters by imaging flow cytometry

图4 PHZ诱导小鼠应激造血模型A：对照组(NS)与实验组(PHZ)的脾脏；B：NS和PHZ组脾脏的质量；C~G：NS和PHZ组的外周血象数据；H：NS和PHZ组骨髓和脾脏收获的细胞总数；I：NS和PHZ组骨髓和脾脏收获的有核红细胞总数；J：NS和PHZ组富集后骨髓和脾脏的有核红细胞总数；K：NS和PHZ组骨髓和脾脏收获的有核红细胞的圈门。*、**、***分别表示差异在P0.05、P0.01、P0.001水平上具有统计学意义；ns表示无统计学意义。

Fig. 4 PHZ-induced stress erythropoiesis model in mice

图5 应激造血状态下骨髓和脾脏中的EBI的细胞组成A：骨髓和脾脏中F4/80+的巨噬细胞在富集前后的细胞比例；B：骨髓和脾脏中F4/80+细胞的流式图；C：骨髓和脾脏中Ter119+的细胞在富集前后的细胞比例；D：骨髓和脾脏中Ter119+细胞的流式图；E：骨髓和脾脏中红系各阶段细胞在富集前的比例；F：骨髓和脾脏中红系各阶段细胞在富集前后的流式图；G：骨髓和脾脏中红系各阶段细胞在富集后的比例；H：骨髓和脾脏CD11b+ Gr-1+细胞在富集前后的比例；I：骨髓和脾脏CD11b+ Gr-1low细胞在富集前后的比例；J：骨髓和脾脏CD11b+ Gr-1hi细胞在富集前后的比例；K：骨髓和脾脏中粒/单核细胞在富集前后的流式图；L：骨髓和脾脏B220low细胞在富集前后的比例；M：骨髓和脾脏B220hi细胞在富集前后的比例；N：骨髓和脾脏中B细胞在富集前后的流式图；O：骨髓和脾脏T细胞在富集前后的比例；P：骨髓和脾脏中T细胞在富集前后的流式图。*、**、***分别表示差异在P0.05、P0.01、P0.001水平上具有统计学意义；ns表示无统计学意义。

Fig. 5 Cellular composition of erythroblastic islands in bone marrow and spleen during stress hematopoiesis

图6 成像流式分析应激造血时骨髓与脾脏细胞团的结构差异及细胞组成A：正常骨髓样本圈取明场面积400的群体为R1；B：BM EBI中F4/80+的事件占R1的比例；C：BM EBI中CD71+的事件占R1的比例；D：BM EBI中F4/80+且CD71+的事件占R1的比例；E：BM EBI中CD11b+且CD71+的事件占R1的比例；F：BM EBI中CD11b+且F4/80+的事件占R1的比例；G：BM EBI中B220+且F4/80+的事件占R1的比例；H：BM EBI中各事件占R1的比例；I：BM EBI的影像；J：SP EBI的影像；K：正常脾脏样本圈取明场面积400的群体为R1；L：SP EBI中F4/80+的事件占R1的比例；M：SP EBI中CD71+的事件占R1的比例；N：SP EBI中F4/80+且CD71+的事件占R1的比例；O：SP EBI中CD11b+且CD71+的事件占R1的比例；P：SP EBI中CD11b+且F4/80+的事件占R1的比例；Q：SP EBI中B220+且F4/80+的事件占R1的比例；R：SP EBI中各事件占R1的比例。*、**、***分别表示差异在P0.05、P0.01、P0.001水平上具有统计学意义。

Fig. 6 Structural and compositional characterization of bone marrow and splenic cell clusters by imaging flow cytometry under stress erythropoiesis

图7 稳态与应激相比EBI的细胞组成变化A：骨髓中富集后的EBI的各谱系细胞比例；B：脾脏中富集后的EBI的各谱系细胞比例；C：骨髓中富集后的EBI的红系各阶段细胞比例；D：脾脏中富集后的EBI的红系各阶段细胞比例；E：骨髓EBI的IFC分析；F：脾脏EBI的IFC分析。*、**分别表示差异在P0.05、P0.01水平上具有统计学意义。

Fig. 7 Comparative analysis of erythroblastic island cellular composition under steady-state and stress conditions

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生理和应激状态下骨髓与脾脏EBI的富集与分析

Comparative Analysis of Erythroblastic Island in Bone Marrow Versus Spleen Under Steady-state and Stress Conditions

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