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首页> 外文期刊>BioResearch open access. >Eighteen-Year Cryopreservation Does Not Negatively Affect the Pluripotency of Human Embryos: Evidence from Embryonic Stem Cell Derivation
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Eighteen-Year Cryopreservation Does Not Negatively Affect the Pluripotency of Human Embryos: Evidence from Embryonic Stem Cell Derivation

机译：十八年低温保存不会负面影响人类胚胎的多能性：胚胎干细胞衍生的证据。

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Human embryonic stem (hES) cells are considered to be a potential source for the therapy of human diseases, drug screening, and the study of developmental biology. In the present study, we successfully derived hES cell lines from blastocysts developed from frozen and fresh embryos. Seventeen- to eighteen-year-old frozen embryos were thawed, cultured to the blastocyst stage, and induced to form hES cells using human foreskin fibroblasts. The Chula2.hES cell line and the Chula4.hES and Chula5.hES cell lines were derived from blastocysts developed from frozen and fresh embryos, respectively. The cell lines expressed pluripotent markers, including alkaline phosphatase (AP), Oct3/4, stage-specific embryonic antigen (SSEA)-4, and tumor recognition antigen (TRA)-1-60 and TRA-1-81 as detected with immunocytochemistry. The real-time polymerase chain reaction (RT-PCR) results showed that the cell lines expressed pluripotent genes, including OCT3/4, SOX2, NANOG, UTF, LIN28, REX1, NODAL , and E-Cadherin . In addition, the telomerase activities of the cell lines were higher than in the fibroblast cells. Moreover, the cell lines differentiated into all three germ layers both in vitro and in vivo . The cell lines had distinct identities, as revealed with DNA fingerprinting, and maintained their normal karyotype after a long-term culture. This study is the first to report the successful derivation of hES cell lines in Thailand and that frozen embryos maintained their pluripotency similar to fresh embryos, as shown by the success of hES cell derivation, even after years of cryopreservation. Therefore, embryos from prolonged cryopreservation could be an alternative source for embryonic stem cell research.
机译：人类胚胎干（hES）细胞被认为是治疗人类疾病，药物筛选和发育生物学研究的潜在来源。在本研究中，我们成功地从冷冻和新鲜胚胎发育的胚泡中获得了hES细胞系。解冻17至18岁的冷冻胚胎，培养至胚泡期，并使用人包皮成纤维细胞诱导形成hES细胞。 Chula2.hES细胞系以及Chula4.hES和Chula5.hES细胞系分别来自冷冻和新鲜胚胎发育的胚泡。这些细胞系表达了多能标记，包括用免疫细胞化学检测到的碱性磷酸酶（AP），Oct3 / 4，阶段特异性胚胎抗原（SSEA）-4和肿瘤识别抗原（TRA）-1-60和TRA-1-81。 。实时聚合酶链反应（RT-PCR）结果表明，细胞系表达了多能基因，包括OCT3 / 4，SOX2，NANOG，UTF，LIN28，REX1，NODAL和E-钙黏着蛋白。另外，细胞系的端粒酶活性高于成纤维细胞。而且，细胞系在体外和体内都分化为所有三个胚层。 DNA指纹显示这些细胞系具有不同的身份，经过长期培养后仍保持其正常的核型。这项研究是第一个报道成功在泰国成功扩增hES细胞系的研究，并且冷冻胚胎保持了与新鲜胚胎相似的多能性，这通过hES细胞衍生的成功得以证明，即使经过多年的冷冻保存也是如此。因此，长期冷冻保存的胚胎可能成为胚胎干细胞研究的替代来源。

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