Aberrant gene expression and sexually incompatible genomic imprinting in oocytes derived from XY mouse embryonic stem cells in vitro.
Aberrant gene expression and sexually incompatible genomic imprinting in oocytes derived from XY mouse embryonic stem cells in vitro.
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DOI：
10.1371/journal.pone.0058555
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Miyoshi J
中科院分区：
文献类型：
--
作者：
Nitta M;Imamura M;Inoue Y;Kunitomo Y;Lin ZY;Ogawa T;Yogo K;Ishida-Kitagawa N;Fukunaga N;Okano H;Sato E;Takeya T;Miyoshi J
小鼠胚胎干细胞（ESCs）具有在体内和体外分化为生殖细胞（GCs）的潜能。有趣的是，XY ESC可以在培养中产生雄性和雌性GC，而不管遗传性别如何。最近的研究表明，胚胎干细胞衍生的原始GCs有助于在体内的功能配子发生，然而，在体外分化技术从来没有成功地产生成熟的卵母细胞从胚胎干细胞由于隐源性生长停滞在腔前卵泡发育阶段。为了解决这个问题，建立了一个小鼠胚胎干细胞系，能够有效地产生卵泡样结构（FLS），使用传统的分子生物学方法研究其特性。结果显示，胚胎干细胞衍生的FLS在形态上类似于卵巢的初级至次级卵泡，但从未形成腔;相反，FLS最终在培养中发生异常发育或细胞死亡，或在移植到小鼠肾包膜下时形成畸胎瘤。基因表达分析表明，FLSs缺乏转录后期卵泡发育所必需的基因，包括促性腺激素受体和类固醇生成酶，而其他一些基因在FLSs过表达相比，成人卵巢。参与细胞间相互作用的E-钙粘蛋白也异位表达。值得注意的是，在FLS中的卵母细胞样细胞表现出雄激素基因组印记，这通常是男性GC的指示。虽然FLS不表达雄性GC标记基因，但DNA甲基转移酶Dnmt 3L以异常高的水平表达。此外，性别决定因素的表达是不明确的，在FLSs的男性和女性的决定因素表达微弱。这些数据表明，胚胎干细胞衍生的FLS的发育功能障碍可能是由于异常的基因表达和基因组印记，可能与不确定的性别决定文化。
Mouse embryonic stem cells (ESCs) have the potential to differentiate into germ cells (GCs) in vivo and in vitro. Interestingly, XY ESCs can give rise to both male and female GCs in culture, irrespective of the genetic sex. Recent studies showed that ESC-derived primordial GCs contributed to functional gametogenesis in vivo; however, in vitro differentiation techniques have never succeeded in generating mature oocytes from ESCs due to cryptogenic growth arrest during the preantral follicle stages of development. To address this issue, a mouse ESC line, capable of producing follicle-like structures (FLSs) efficiently, was established to investigate their properties using conventional molecular biological methods. The results revealed that the ESC-derived FLSs were morphologically similar to ovarian primary-to-secondary follicles but never formed an antrum; instead, the FLSs eventually underwent abnormal development or cell death in culture, or formed teratomas when transplanted under the kidney capsule in mice. Gene expression analyses demonstrated that the FLSs lacked transcripts for genes essential to late folliculogenesis, including gonadotropin receptors and steroidogenic enzymes, whereas some other genes were overexpressed in FLSs compared to the adult ovary. The E-Cadherin protein, which is involved in cell-to-cell interactions, was also expressed ectopically. Remarkably, it was seen that oocyte-like cells in the FLSs exhibited androgenetic genomic imprinting, which is ordinarily indicative of male GCs. Although the FLSs did not express male GC marker genes, the DNA methyltransferase, Dnmt3L, was expressed at an abnormally high level. Furthermore, the expression of sex determination factors was ambiguous in FLSs as both male and female determinants were expressed weakly. These data suggest that the developmental dysfunction of the ESC-derived FLSs may be attributable to aberrant gene expression and genomic imprinting, possibly associated with uncertain sex determination in culture.
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