Overexpression of BCL2 enhances survival of human embryonic stem cells during stress and obviates the requirement for serum factors
Overexpression of BCL2 enhances survival of human embryonic stem cells during stress and obviates the requirement for serum factors
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DOI：
10.1073/pnas.1019047108
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发表时间：
2011-02-22
影响因子：
11.1
通讯作者：
Weissman, Irving L.
中科院分区：
文献类型：
--
作者：
Ardehali, Reza;Inlay, Matthew A.;Weissman, Irving L.
多能干细胞作为研究和治疗工具的前景在一定程度上因在培养物中生成和维持这些细胞的技术挑战而受到削弱。人胚胎干细胞 (hESC) 对培养条件极其敏感，需要生长因子以及细胞-细胞和细胞-基质相互作用的持续信号传导，以防止细胞凋亡、衰老和分化。我们实验室之前的工作表明，小鼠胚胎干细胞中促存活基因 BCL2 的过度表达超越了维持 mESC 培养所需的血清因子和饲养细胞的需求。为了确定这种促存活基因是否可以类似地保护 hESC，我们生成了组成型或诱导型表达 BCL2 的 hESC 系。我们发现 BCL2 过表达显着减少解离诱导的细胞凋亡，导致分选的单细胞集落形成增强，并增强胚状体形成。此外，BCL2-hESC 在没有血清的情况下表现出正常生长，但需要碱性成纤维细胞生长因子才能保持未分化。此外，它们保持多能性标记，在体内形成畸胎瘤，并分化成所有三个胚层。我们的数据表明，BCL2 信号通路在抑制 hESC 凋亡中发挥重要作用，因此其在 hESC 中的过度表达既可以通过抵抗细胞凋亡而在应激条件下提供生存益处，又可以消除对血清或饲养层维持的需要。
The promise of pluripotent stem cells as a research and therapeutic tool is partly undermined by the technical challenges of generating and maintaining these cells in culture. Human embryonic stem cells (hESCs) are exquisitely sensitive to culture conditions, and require constant signaling by growth factors and cell-cell and cell-matrix interactions to prevent apoptosis, senescence, and differentiation. Previous work from our laboratory demonstrated that overexpression of the prosurvival gene BCL2 in mouse embryonic stem cells overrode the requirement of serum factors and feeder cells to maintain mESCs in culture. To determine whether this prosurvival gene could similarly protect hESCs, we generated hESC lines that constitutively or inducibly express BCL2. We find that BCL2 overexpression significantly decreases dissociation-induced apoptosis, resulting in enhanced colony formation from sorted single cells, and enhanced embryoid body formation. In addition, BCL2-hESCs exhibit normal growth in the absence of serum, but require basic fibroblast growth factor to remain undifferentiated. Furthermore, they maintain their pluripotency markers, form teratomas in vivo, and differentiate into all three germ layers. Our data suggest that the BCL2 signaling pathway plays an important role in inhibiting hESC apoptosis, such that its overexpression in hESCs offers both a survival benefit in conditions of stress by resisting apoptosis and obviates the requirement for serum or a feeder layer for maintenance.