Directed Differentiation of Human Embryonic Stem Cells into Prostate Organoids In Vitro and its Perturbation by Low-Dose Bisphenol A Exposure.
Directed Differentiation of Human Embryonic Stem Cells into Prostate Organoids In Vitro and its Perturbation by Low-Dose Bisphenol A Exposure.
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DOI：
10.1371/journal.pone.0133238
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Prins GS
中科院分区：
文献类型：
--
作者：
Calderon-Gierszal EL;Prins GS
使用啮齿动物和成年人类前列腺干细胞-祖细胞模型的研究表明，发育过程中暴露于内分泌干扰物双酚A(BPA)会随着年龄的增长而易于发生前列腺癌。目前尚不清楚人类胚胎前列腺在其自然发育窗口期是否同样容易受到BPA的影响。为了解决这一未得到满足的需求，我们在此报告了一个开创性的体外人类前列腺发育模型的构建，以研究双酚A的影响。人类胚胎干细胞(HESC)在空间系统中定向分化为前列腺器官是通过对生长因子和类固醇的精确时间控制完成的。激活素诱导的最终内胚层通过联合暴露于WNT10B和FGF10而达到前列腺规格。Matrigel在含有R-Spinin-1、Noggin、EGF、维甲酸和睾酮的培养液中培养20-30天，就足以发育成熟的前列腺器官。免疫荧光和基因表达分析证实，有机化合物具有人类前列腺的细胞分化和功能特性。在分化培养过程中，1 nM或10 nM双酚A以剂量依赖的方式干扰早期形态发生，1 nM双酚A增加，10 nM双酚A减少分枝结构的形成。虽然分支结构向成熟类器官的分化似乎在很大程度上不受BPA暴露的影响，但类干细胞数量增加，表现为没有正确进入谱系承诺的焦点干细胞巢，而不是在正常分化结构中发现的罕见的分离干细胞。这些发现首次提供了直接证据，表明低剂量双酚A暴露以hESC为靶点，并扰乱胚胎细胞向人类前列腺器官分化的形态发生，表明发育中的人类前列腺可能容易受到宫内双酚A暴露的破坏。
Studies using rodent and adult human prostate stem-progenitor cell models suggest that developmental exposure to the endocrine disruptor Bisphenol-A (BPA) can predispose to prostate carcinogenesis with aging. Unknown at present is whether the embryonic human prostate is equally susceptible to BPA during its natural developmental window. To address this unmet need, we herein report the construction of a pioneer in vitro human prostate developmental model to study the effects of BPA. The directed differentiation of human embryonic stem cells (hESC) into prostatic organoids in a spatial system was accomplished with precise temporal control of growth factors and steroids. Activin-induced definitive endoderm was driven to prostate specification by combined exposure to WNT10B and FGF10. Matrigel culture for 20–30 days in medium containing R-Spondin-1, Noggin, EGF, retinoic acid and testosterone was sufficient for mature prostate organoid development. Immunofluorescence and gene expression analysis confirmed that organoids exhibited cytodifferentiation and functional properties of the human prostate. Exposure to 1 nM or 10 nM BPA throughout differentiation culture disturbed early morphogenesis in a dose-dependent manner with 1 nM BPA increasing and 10 nM BPA reducing the number of branched structures formed. While differentiation of branched structures to mature organoids seemed largely unaffected by BPA exposure, the stem-like cell population increased, appearing as focal stem cell nests that have not properly entered lineage commitment rather than the rare isolated stem cells found in normally differentiated structures. These findings provide the first direct evidence that low-dose BPA exposure targets hESC and perturbs morphogenesis as the embryonic cells differentiate towards human prostate organoids, suggesting that the developing human prostate may be susceptible to disruption by in utero BPA exposures.