Goumans M J, Ward-van Oostwaard D, Wianny F, Savatier P, Zwijsen A, Mummery C
Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Utrecht, The Netherlands.
Differentiation. 1998 Jul;63(3):101-13. doi: 10.1046/j.1432-0436.1998.6330101.x.
Embryonic stem (ES) cells are resistant to transforming growth factor beta (TGF beta). We have shown previously that they lack type-II binding receptors (T beta RII) and in this respect resemble the inner cell mass and ectoderm cells of mouse embryos 4.5-7.5 days post coitum (dpc); they do however express type-I (alk-5) signalling receptors. Here we show that in contrast to several tumour cell lines, stable transfection of wtT beta RII is not sufficient for ES cells to become biologically sensitive to TGF beta. We analysed the expression of several down-stream molecules known to be involved in TGF beta signalling (Smads) and TGF beta-mediated cell cycle regulation (cyclins D) during the differentiation of control and wtT beta RII-expressing ES cells and showed that upregulation of these molecules correlated with (i) an increase in plasminogen activator inhibitor-1 (PAI-1) synthesis and (ii) growth inhibition, following addition of TGF beta 1. These TGF beta responses were reduced in an ES cell line expressing a dominant negative (truncated) T beta RII (delta T beta RII). The differentiation pattern of control and wtT beta RII-expressing ES cells was indistinguishable in monolayer culture and as embryoid bodies, but in delta T beta RII ES cells, the capacity to form mesodermal derivatives in monolayer cultures in response to the addition of retinoic acid (RA) and removal of leukemia inhibitory factor (LIF) was lost, and only endoderm-like cells formed. The T beta RII and delta T beta RII ES cells were, however, both distinguishable from control ES cells when allowed to differentiate in chimaeric embryos following aggregation with morula-stage hosts. Conceptuses containing mutant cells, recovered from pseudopregnant females at the equivalent of 9.5 dpc, exhibited highly defective yolk sac development; most strikingly, no blood vessels were present and in addition the yolk sacs with derivatives of ES cells containing wtT beta RII were blistered and lacked haematopoietic cells. The implications for understanding TGF beta signalling in early mouse development are discussed.
胚胎干细胞（ES细胞）对转化生长因子β（TGFβ）具有抗性。我们之前已经表明，它们缺乏II型结合受体（TβRII），在这方面类似于交配后4.5 - 7.5天（dpc）的小鼠胚胎的内细胞团和外胚层细胞；然而，它们确实表达I型（alk - 5）信号受体。在这里我们表明，与几种肿瘤细胞系不同，野生型TβRII的稳定转染不足以使ES细胞对TGFβ产生生物学敏感性。我们分析了在对照和表达野生型TβRII的ES细胞分化过程中，已知参与TGFβ信号传导（Smads）和TGFβ介导的细胞周期调控（细胞周期蛋白D）的几种下游分子的表达，并表明这些分子的上调与（i）纤溶酶原激活物抑制剂 - 1（PAI - 1）合成增加以及（ii）添加TGFβ1后生长抑制相关。在表达显性负性（截短）TβRII（ΔTβRII）的ES细胞系中，这些TGFβ反应减弱。对照和表达野生型TβRII的ES细胞在单层培养和形成胚状体时的分化模式无法区分，但在ΔTβRII ES细胞中，响应视黄酸（RA）添加和白血病抑制因子（LIF）去除，在单层培养中形成中胚层衍生物的能力丧失，仅形成内胚层样细胞。然而，当与桑椹胚期宿主聚集后在嵌合胚胎中分化时，TβRII和ΔTβRII ES细胞都与对照ES细胞不同。在相当于9.5 dpc时从假孕雌性中回收的含有突变细胞的概念胚胎，其卵黄囊发育存在严重缺陷；最显著的是，没有血管存在，此外，含有野生型TβRII的ES细胞衍生物的卵黄囊出现水泡样且缺乏造血细胞。本文讨论了对理解小鼠早期发育中TGFβ信号传导的意义。
