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一种改良培养基可增加囊胚形成率以及从质量较差胚胎中获得人胚胎干细胞的效率。

A modified culture medium increases blastocyst formation and the efficiency of human embryonic stem cell derivation from poor-quality embryos.

作者信息

FAN Yong, LUO Yumei, CHEN Xinjie, SUN Xiaofang

机构信息

Institute of Gynecology and Obstetrics, The Third Affiliated Hospital of Guangzhou Medical College, Guangzhou, Guangdong, China.

出版信息

J Reprod Dev. 2010 Oct;56(5):533-9. doi: 10.1262/jrd.09-225m. Epub 2010 Jul 20.

Abstract

Human embryonic stem cells (HESCs) are defined as self-renewing cells that retain their ability to differentiate into all cell types of the body. They have enormous potential in medical applications and as a model for early human development. There is a need for derivation of new HESC lines to meet emerging requirements for their use in cell replacement therapies, disease modeling, and basic research. Here, we describe a modified culture medium containing human recombinant leukemia inhibitory factor and human basic fibroblast growth factor that significantly increases the number of human blastocysts formed and their quality, as well as the efficiency of HESC derivation from poor-quality embryos. Culturing poor-quality embryos in modified medium resulted in a two-fold increase in the blastocyst formation rate and a seven-fold increase over the derivation efficiency in conventional medium. We derived 15 HESC lines from poor-quality embryos cultured in modified culture medium and two HESC lines from quality embryos cultured in conventional culture medium. All cell lines shared typical human pluripotent stem cell features including similar morphology, normal karyotypes, expression of alkaline phosphatase, pluripotency genes, such as Oct4, and cell surface markers (SSEA-4, TRA-1-60, TRA-1-81), the ability to form teratomas in SCID mice, and the ability to differentiate into cells of three embryonic germ layers in vitro. Our data suggest that poor-quality embryos that have reached the blastocyst stage in our modified culture medium are a robust source for normal HESC line derivation.

摘要

人胚胎干细胞(HESCs)被定义为能够自我更新且保留分化为身体所有细胞类型能力的细胞。它们在医学应用以及作为早期人类发育模型方面具有巨大潜力。为满足细胞替代疗法、疾病建模和基础研究中对其使用的新需求,需要衍生新的HESC系。在此,我们描述了一种改良培养基,其含有重组人白血病抑制因子和人碱性成纤维细胞生长因子,可显著增加形成的人囊胚数量及其质量,以及从质量较差胚胎中衍生HESC的效率。在改良培养基中培养质量较差的胚胎,囊胚形成率提高了两倍,衍生效率比传统培养基提高了七倍。我们从在改良培养基中培养的质量较差胚胎中衍生出15个HESC系,从在传统培养基中培养的优质胚胎中衍生出2个HESC系。所有细胞系都具有典型的人多能干细胞特征,包括相似的形态、正常的核型、碱性磷酸酶的表达、多能性基因(如Oct4)以及细胞表面标志物(SSEA - 4、TRA - 1 - 60、TRA - 1 - 81),能够在SCID小鼠中形成畸胎瘤,并且能够在体外分化为三个胚胎胚层的细胞。我们的数据表明,在我们的改良培养基中发育到囊胚阶段的质量较差胚胎是正常HESC系衍生的可靠来源。

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