Differentiation of lymphatic endothelial cells from embryonic stem cells on OP9 stromal cells
Differentiation of lymphatic endothelial cells from embryonic stem cells on OP9 stromal cells
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DOI：
10.1161/01.atv.0000225770.57219.b0
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发表时间：
2006-09-01
影响因子：
8.7
通讯作者：
Yamashita, Jun K.
中科院分区：
文献类型：
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作者：
Kono, Tomoya;Kubo, Hajime;Yamashita, Jun K.
目的：血管内皮生长因子C （VEGF-C）和VEGF受体-3 （VEGFR-3）的发现已经开始提供对淋巴管生成的分子机制的理解。同源盒基因prox1已被证明可以从血液内皮细胞中指定淋巴内皮细胞（ECs）。我们利用胚胎干细胞（ES）研究淋巴EC （LEC）分化的过程。方法和结果胚胎干细胞衍生的-VEGFR-2（+）细胞在OP9基质细胞上分化为LECs，通过表达prox1、VEGFR-3和另一种淋巴标记物podoplanin来定义。VEGFR-2（+）细胞产生LYVE-1（+）胚胎ECs，第1天prox1为阴性，第3天变为prox1(+) LECs。VEGFR-3-Fc或Tie2-Fc，隔离VEGF-C或血管生成素1 (Ang1)，抑制了OP9细胞上LECs集落的形成。然而，在胶原包被培养皿中，在VEGFR-2（+）细胞中加入VEGF- c和Ang1并与VEGF联合培养，均未能诱导出LECs。在多聚甲醛固定的OP9细胞上，条件培养基完全复制了OP9细胞的lec诱导活性。结论：我们成功地从ES细胞中分化出了LEC，并揭示了VEGF-C、Ang1等未知因子对LEC分化的要求。
Objectives - The discovery of vascular endothelial growth factor C (VEGF-C) and VEGF receptor-3 (VEGFR-3) has started to provide an understanding of the molecular mechanisms of lymphangiogenesis. The homeobox gene prox1 has been proven to specify lymphatic endothelial cells (ECs) from blood ECs. We investigated the process of lymphatic EC (LEC) differentiation using embryonic stem (ES) cells.Methods and Results -VEGFR-2(+) cells derived from ES cells differentiated into LECs at day 3 on OP9 stromal cells defined by the expression of prox1, VEGFR-3, and another lymphatic marker podoplanin. VEGFR-2(+) cells gave rise to LYVE-1(+) embryonic ECs, which were negative for prox1 on day 1 but turned to prox1(+) LECs by day 3. VEGFR-3-Fc or Tie2-Fc, sequestering VEGF-C or angiopoietin1 (Ang1), suppressed colony formation of LECs on OP9 cells. However, addition of VEGF-C and Ang1 in combination with VEGF to the culture of VEGFR-2(+) cells on collagen-coated dishes failed to induce LECs. LEC-inducing activity of OP9 cells was fully reproduced on paraformaldehyde-fixed OP9 cells with the conditioned medium.Conclusion - We succeeded in differentiating LECs from ES cells and revealed the requirements of VEGF-C, Ang1, and other unknown factors for LEC differentiation.