Chromosome 7 and 19 trisomy in cultured human neural progenitor cells.
Chromosome 7 and 19 trisomy in cultured human neural progenitor cells.
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DOI：
10.1371/journal.pone.0007630
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发表时间：
2009-10-29
期刊：
影响因子：
3.7
通讯作者：
Svendsen CN
中科院分区：
文献类型：
--
作者：
Sareen D;McMillan E;Ebert AD;Shelley BC;Johnson JA;Meisner LF;Svendsen CN
干细胞扩增和分化是新兴细胞治疗技术的基础。人类神经祖细胞（hNPC）的潜在应用范围广泛，但正常的细胞遗传学特征对于避免临床试验中肿瘤形成的风险非常重要。 FDA 批准的临床试验正在计划和进行，将 hNPC 移植到大脑或脊髓中，以治疗各种神经退行性疾病。尽管已知人类胚胎干细胞 (hESC) 会反复出现涉及 12 和 17 的染色体异常，但尚无研究揭示培养的 hNPC 中存在染色体异常。因此，我们研究了 21 个独立的胎儿来源 hNPC 系中经常发生的染色体异常以及触发此类畸变的可能机制。虽然大多数 hNPC 系的核型正常，但 G 带核型分析和荧光原位杂交 (FISH) 分析显示，分别在 24% 和 5% 的系中出现了 7 三体性 (hNPC+7) 和 19 三体性 (hNPC+19)。一旦检测到，随后的传代揭示了三体性 hNPC 的新兴优势。 DNA 微阵列和免疫印迹分析表明 hNPC+7 和 hNPC+19 细胞中表皮生长因子受体 (EGFR) 过度表达。我们使用各自的免疫细胞化学标记物观察到 hNPC+7 和 hNPC+19 中端粒酶 (hTERT) 水平更高、增殖 (Ki67) 增加、存活 (TUNEL) 和神经发生 (βIII-微管蛋白) 增加。然而，三体系在 50-60 次群体倍增后经历了复制性衰老，并且从未表现出肿瘤性变化。虽然hNPC+7和hNPC+19在异种移植到大鼠纹状体后存活得更好，但它们没有形成恶性肿瘤。最后，EGF 剥夺引发了二倍体 hNPC 系中 7 三体细胞的选择。我们报告说，hNPC 在长期细胞培养中容易受到 7 号和 19 号染色体三体积累的影响。这些结果表明，微环境线索是选择特定 hNPC 非整倍体的强大因素，其中 7 号染色体三体性是最常见的。鉴于美国正在进行或计划进行许多基于干细胞的临床试验，并且 PLoS Medicine 最近的一份报告显示了移植过多细胞的危险，这些数据证实了在将 hNPC（胎儿或 hESC 衍生的）用于临床或基础科学应用之前需要对其进行仔细的细胞遗传学评估。
Stem cell expansion and differentiation is the foundation of emerging cell therapy technologies. The potential applications of human neural progenitor cells (hNPCs) are wide ranging, but a normal cytogenetic profile is important to avoid the risk of tumor formation in clinical trials. FDA approved clinical trials are being planned and conducted for hNPC transplantation into the brain or spinal cord for various neurodegenerative disorders. Although human embryonic stem cells (hESCs) are known to show recurrent chromosomal abnormalities involving 12 and 17, no studies have revealed chromosomal abnormalities in cultured hNPCs. Therefore, we investigated frequently occurring chromosomal abnormalities in 21 independent fetal-derived hNPC lines and the possible mechanisms triggering such aberrations. While most hNPC lines were karyotypically normal, G-band karyotyping and fluorescent in situ hybridization (FISH) analyses revealed the emergence of trisomy 7 (hNPC+7) and trisomy 19 (hNPC+19), in 24% and 5% of the lines, respectively. Once detected, subsequent passaging revealed emerging dominance of trisomy hNPCs. DNA microarray and immunoblotting analyses demonstrate epidermal growth factor receptor (EGFR) overexpression in hNPC+7 and hNPC+19 cells. We observed greater levels of telomerase (hTERT), increased proliferation (Ki67), survival (TUNEL), and neurogenesis (βIII-tubulin) in hNPC+7 and hNPC+19, using respective immunocytochemical markers. However, the trisomy lines underwent replicative senescence after 50–60 population doublings and never showed neoplastic changes. Although hNPC+7 and hNPC+19 survived better after xenotransplantation into the rat striatum, they did not form malignant tumors. Finally, EGF deprivation triggered a selection of trisomy 7 cells in a diploid hNPC line. We report that hNPCs are susceptible to accumulation of chromosome 7 and 19 trisomy in long-term cell culture. These results suggest that micro-environmental cues are powerful factors in the selection of specific hNPC aneuploidies, with trisomy of chromosome 7 being the most common. Given that a number of stem cell based clinical trials are being conducted or planned in USA and a recent report in PLoS Medicine showing the dangers of grafting an inordinate number of cells, these data substantiate the need for careful cytogenetic evaluation of hNPCs (fetal or hESC-derived) before their use in clinical or basic science applications.
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