Ebrahimi Saman, Shams Alireza, Maghami Parvaneh, Hekmat Azadeh
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
Department of Anatomy, School of Medicine, Alborz University of Medical Sciences, Karaj, Iran.
Cell J. 2022 Aug 28;24(8):458-464. doi: 10.22074/cellj.2022.8303.
Objective: Primordial germ cell (PGCs) lines are a source of a highly specialized type of cells, characteristically oocytes,
during female germline development in vivo. The oocyte growth begins in the transition from the primary follicle. It is
associated with dynamic changes in gene expression, but the gene-regulating signals and transcription factors that control oocyte growth remain unknown. We aim to investigate the differentiation potential of mouse bone marrow mesenchymal stem cells (mMSCs) into female germ-like cells by testing several signals and transcription factors.
Materials and Methods: In this experimental study, mMSCs were extracted from mice femur bone using the flushing
technique. The cluster-differentiation (CD) of superficial mesenchymal markers was determined with flow cytometric analysis. We applied a set of transcription factors including retinoic acid (RA), titanium nanotubes (TNTs), and fibrin such as TNT-coated fibrin (F+TNT) formation and (RA+F+TNT) induction, and investigated the changes in gene, MVH/ DDX4, expression and functional screening using an in vitro mouse oocyte development condition. Germ cell markers expression, (MVH / DDX4), was analyzed with Immunocytochemistry staining, quantitative transcription-polymerase chain reaction (RT-qPCR) analysis, and Western blots.
Results: The expression of CD was confirmed by flow cytometry. The phase determination of the TNTs and F+TNT were confirmed using x-ray diffraction (XRD) and scanning electron microscope (SEM), respectively. Remarkably, applying these transcription factors quickly induced pluripotent stem cells into oocyte-like cells that were sufficient to generate female germlike cells, growth, and maturation from mMSCs differentiation. These transcription factors formed oocyte-like cells specification of stem cells, epigenetic reprogramming, or meiosis and indicate that oocyte meiosis initiation and oocyte growth are not separable from the previous epigenetic reprogramming in stem cells in vitro.
Conclusion: Results suggested several transcription factors may apply for arranging oocyte-like cell growth and supplies an alternative source of in vitro maturation (IVM).
目的：原始生殖细胞（PGCs）系是一种高度特化的细胞来源，在体内雌性生殖系发育过程中典型地表现为卵母细胞。卵母细胞生长始于从初级卵泡的转变。它与基因表达的动态变化相关，但控制卵母细胞生长的基因调控信号和转录因子仍不清楚。我们旨在通过测试几种信号和转录因子来研究小鼠骨髓间充质干细胞（mMSCs）向雌性生殖样细胞的分化潜能。
材料和方法：在本实验研究中，使用冲洗技术从小鼠股骨中提取mMSCs。通过流式细胞术分析确定表面间充质标志物的簇分化（CD）。我们应用了一组转录因子，包括视黄酸（RA）、钛纳米管（TNTs）和纤维蛋白，如TNT包被的纤维蛋白（F + TNT）形成和（RA + F + TNT）诱导，并使用体外小鼠卵母细胞发育条件研究基因、MVH / DDX4表达的变化和功能筛选。通过免疫细胞化学染色、定量转录 - 聚合酶链反应（RT - qPCR）分析和蛋白质印迹分析生殖细胞标志物（MVH / DDX4）的表达。
结果：通过流式细胞术确认了CD的表达。分别使用X射线衍射（XRD）和扫描电子显微镜（SEM）确认了TNTs和F + TNT的相测定。值得注意的是，应用这些转录因子可迅速将多能干细胞诱导为卵母细胞样细胞，这些细胞足以从mMSCs分化中产生雌性生殖样细胞、生长和成熟。这些转录因子形成了干细胞的卵母细胞样细胞特化、表观遗传重编程或减数分裂，并表明卵母细胞减数分裂起始和卵母细胞生长在体外干细胞中与先前的表观遗传重编程不可分离。
结论：结果表明几种转录因子可能适用于安排卵母细胞样细胞的生长，并提供体外成熟（IVM）的替代来源。
