Enver Tariq, Soneji Shamit, Joshi Chirag, Brown John, Iborra Francisco, Orntoft Torben, Thykjaer Thomas, Maltby Edna, Smith Kath, Abu Dawud Raed, Jones Mark, Matin Maryam, Gokhale Paul, Draper Jonathan, Andrews Peter W
MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, UK.
Hum Mol Genet. 2005 Nov 1;14(21):3129-40. doi: 10.1093/hmg/ddi345. Epub 2005 Sep 13.
Human embryonic stem cell (HESC) lines vary in their characteristics and behaviour not only because they are derived from genetically outbred populations, but also because they may undergo progressive adaptation upon long-term culture in vitro. Such adaptation may reflect selection of variants with altered propensity for survival and retention of an undifferentiated phenotype. Elucidating the mechanisms involved will be important for understanding normal self-renewal and commitment to differentiation and for validating the safety of HESC-based therapy. We have investigated this process of adaptation at the cellular and molecular levels through a comparison of early passage (normal) and late passage (adapted) sublines of a single HESC line, H7. To account for spontaneous differentiation that occurs in HESC cultures, we sorted cells for SSEA3, which marks undifferentiated HESC. We show that the gene expression programmes of the adapted cells partially reflected their aberrant karyotype, but also resulted from a failure in X-inactivation, emphasizing the importance in adaptation of karyotypically silent epigenetic changes. On the basis of growth potential, ability to re-initiate ES cultures and global transcription profiles, we propose a cellular differentiation hierarchy for maintenance cultures of HESC: normal SSEA3+ cells represent pluripotent stem cells. Normal SSEA3- cells have exited this compartment, but retain multilineage differentiation potential. However, adapted SSEA3+ and SSEA3- cells co-segregate within the stem cell territory, implying that adaptation reflects an alteration in the balance between self-renewal and differentiation. As this balance is also an essential feature of cancer, the mechanisms of culture adaptation may mirror those of oncogenesis and tumour progression.
人类胚胎干细胞（HESC）系在其特征和行为上存在差异，这不仅是因为它们源自基因杂合群体，还因为它们在体外长期培养时可能会发生渐进性适应。这种适应可能反映了具有改变的存活倾向和未分化表型保留能力的变体的选择。阐明其中涉及的机制对于理解正常的自我更新和分化承诺以及验证基于HESC的治疗的安全性至关重要。我们通过比较单个HESC系H7的早期传代（正常）和晚期传代（适应）亚系，在细胞和分子水平上研究了这种适应过程。为了解释HESC培养物中发生的自发分化，我们对标记未分化HESC的SSEA3进行细胞分选。我们表明，适应细胞的基因表达程序部分反映了它们异常的核型，但也源于X染色体失活失败，强调了核型沉默的表观遗传变化在适应中的重要性。基于生长潜力、重新启动ES培养的能力和全局转录谱，我们提出了HESC维持培养的细胞分化层次结构：正常的SSEA3 +细胞代表多能干细胞。正常的SSEA3 -细胞已离开这个区室，但保留多谱系分化潜力。然而，适应的SSEA3 +和SSEA3 -细胞在干细胞区域内共同分离，这意味着适应反映了自我更新和分化之间平衡的改变。由于这种平衡也是癌症的一个基本特征，培养适应的机制可能反映了肿瘤发生和肿瘤进展的机制。
