Department of Cell Biology, Institute of Basic Medical Sciences, Beijing, China.
Haematologica. 2010 Jun;95(6):875-83. doi: 10.3324/haematol.2009.014241. Epub 2009 Dec 8.
The hemangioblast is a bi-potential precursor cell with the capacity to differentiate into hematopoietic and vascular cells. In mouse E7.0-7.5 embryos, the hemangioblast can be identified by a clonal blast colony-forming cell (BL-CFC) assay or single cell OP9 co-culture. However, the ontogeny of the hemangioblast in mid-gestation embryos is poorly defined.
The BL-CFC assay and the OP9 system were combined to illustrate the hemangioblast with lymphomyeloid and vascular potential in the mouse aorta-gonad-mesonephros region. The colony-forming assay, reverse transcriptase polymerase chain reaction analysis, immunostaining and flow cytometry were used to identify the hematopoietic potential, and Matrigel- or OP9-based methods were employed to evaluate endothelial progenitor activity.
Functionally, the aorta-gonad-mesonephros-derived BL-CFC produced erythroid/myeloid progenitors, CD19(+) B lymphocytes, and CD3(+)TCRbeta(+) T lymphocytes. Meanwhile, the BL-CFC-derived adherent cells generated CD31(+) tube-like structures on OP9 stromal cells, validating the endothelial progenitor potential. The aorta-gonad-mesonephros-derived hemangioblast was greatly enriched in CD31(+), endomucin(+) and CD105(+) subpopulations, which collectively pinpoints the endothelial layer as the main location. Interestingly, the BL-CFC was not detected in yolk sac, placenta, fetal liver or embryonic circulation. Screening of candidate cytokines revealed that interleukin-3 was remarkable in expanding the BL-CFC in a dose-dependent manner through the JAK2/STAT5 and MAPK/ERK pathways. Neutralizing interleukin-3 in the aorta-gonad-mesonephros region resulted in reduced numbers of BL-CFC, indicating the physiological requirement for this cytokine. Both hematopoietic and endothelial differentiation potential were significantly increased in interleukin-3-treated BL-CFC, suggesting a persistent positive influence. Intriguingly, interleukin-3 markedly amplified primitive erythroid and macrophage precursors in E7.5 embryos. Quantitative polymerase chain reaction analysis demonstrated declined Flk-1 and elevated Scl and von Willebrand factor transcription upon interleukin-3 stimulation, indicating accelerated hemangiopoiesis.
The hemangioblast with lymphomyeloid potential is one of the precursors of definitive hematopoiesis in the mouse aorta-gonad-mesonephros region. Interleukin-3 has a regulatory role with regards to both the number and capacity of the dual-potential hemangioblast.
血岛细胞是一种具有双重潜能的前体细胞，能够分化为造血细胞和血管细胞。在小鼠 E7.0-7.5 胚胎中，可以通过克隆胚集落形成细胞（BL-CFC）测定或单细胞 OP9 共培养来鉴定血岛细胞。然而，中孕期胚胎中血岛细胞的发生情况尚不清楚。
将 BL-CFC 测定法和 OP9 系统相结合，以说明在小鼠主动脉-性腺-中肾区具有淋巴骨髓和血管潜能的血岛细胞。集落形成测定、逆转录聚合酶链反应分析、免疫染色和流式细胞术用于鉴定造血潜能，Matrigel 或 OP9 方法用于评估内皮祖细胞活性。
从主动脉-性腺-中肾区分离的 BL-CFC 具有产生红系/髓系祖细胞、CD19(+)B 淋巴细胞和 CD3(+)TCRβ(+)T 淋巴细胞的功能。同时，BL-CFC 衍生的贴壁细胞在 OP9 基质细胞上产生 CD31(+)管状结构，验证了内皮祖细胞的潜能。从主动脉-性腺-中肾区分离的血岛细胞在 CD31(+)、内粘蛋白(+)和 CD105(+)亚群中得到了极大的富集，这共同指出了内皮层是主要位置。有趣的是，BL-CFC 未在卵黄囊、胎盘、胎肝或胚胎循环中检测到。对候选细胞因子的筛选表明，白细胞介素-3 通过 JAK2/STAT5 和 MAPK/ERK 途径以剂量依赖性方式显著扩增 BL-CFC。在主动脉-性腺-中肾区中和白细胞介素-3 会导致 BL-CFC 数量减少，表明该细胞因子具有生理需求。白细胞介素-3 处理后的 BL-CFC 的造血和内皮分化潜能均显著增加，表明存在持续的正向影响。有趣的是，白细胞介素-3 显著扩增了 E7.5 胚胎中的原始红细胞和巨噬细胞前体。定量聚合酶链反应分析表明，白细胞介素-3 刺激后 Flk-1 转录下调，Scl 和 von Willebrand 因子转录上调，提示加速了造血发生。
具有淋巴骨髓潜能的血岛细胞是小鼠主动脉-性腺-中肾区中确定造血的前体细胞之一。白细胞介素-3 对双潜能血岛细胞的数量和能力具有调节作用。
