Human bone marrow-derived mesenchymal stem cells do not undergo transformation after long-term In vitro culture and do not exhibit telomere maintenance mechanisms
Human bone marrow-derived mesenchymal stem cells do not undergo transformation after long-term In vitro culture and do not exhibit telomere maintenance mechanisms
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DOI：
10.1158/0008-5472.can-06-4690
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发表时间：
2007-10-01
期刊：
影响因子：
11.2
通讯作者：
Locatelli, Franco
中科院分区：
文献类型：
--
作者：
Bernardo, Maria Ester;Zaffaroni, Nadia;Locatelli, Franco
对间充质干细胞（MSC）生物学认识的重大进步为其临床应用开辟了道路。然而，人们对间充质干细胞发生恶性转化的可能性产生了担忧。我们研究了人骨髓（BM）来源的间充质干细胞在不同体外培养时间点的转化易感性。从10名健康供体的骨髓中分离出间充质干细胞，并在体外进行培养，直至细胞衰老或传代至第25代（P25）。在终止培养前，对处于衰老阶段的间充质干细胞密切监测8至12周。在长期培养前后，通过阵列比较基因组杂交（array - CGH）、常规核型分析和亚端粒荧光原位杂交分析对间充质干细胞进行基因特征研究。在不同传代时检测间充质干细胞的端粒酶活性、人端粒酶逆转录酶（hTERT）转录本以及端粒替代延长（ALT）机制的表达。注意到供体之间在增殖能力和间充质干细胞寿命方面存在巨大差异。在10名供体中的8名，间充质干细胞的增殖能力逐渐下降直至衰老。在其余2个间充质干细胞样本中，培养在第25代时终止以进行数据分析。阵列比较基因组杂交和细胞遗传学分析表明，体外扩增的间充质干细胞未显示染色体异常。在所有检测的培养物中均未检测到端粒酶活性和hTERT转录本，并且在培养期间端粒缩短。在测试的间充质干细胞中未发现端粒替代延长现象。骨髓来源的间充质干细胞可在体外安全扩增，不易发生恶性转化，因此这些细胞适合用于细胞治疗方法。
Significant improvement in the understanding of mesenchymal stem cell (MSC) biology has opened the way to their clinical use. However, concerns regarding the possibility that MSCs undergo malignant transformation have been raised. We investigated the susceptibility to transformation of human bone marrow (BM)-derived MSCs at different in vitro culture time points. MSCs were isolated from BM of 10 healthy donors and propagated in vitro until reaching either senescence or passage (P) 25. MSCs in the senescence phase were closely monitored for 8 to 12 weeks before interrupting the cultures. The genetic characterization of MSCs was investigated through array-comparative genomic hybridization (array-CGH), conventional karyotyping, and subtelomeric fluorescent in situ hybridization analysis both before and after prolonged culture. MSCs were tested for the expression of telomerase activity, human telomerase reverse transcriptase (hTERT) transcripts, and alternative lengthening of telomere (ALT) mechanism at different passages. A huge variability in terms of proliferative capacity and MSCs life span was noted between donors. In eight of 10 donors, MSCs displayed a progressive decrease in proliferative capacity until reaching senescence. In the remaining two MSC samples, the cultures were interrupted at P25 to pursue data analysis. Array-CGH and cytogenetic analyses showed that MSCs expanded in vitro did not show chromosomal abnormalities. Telomerase activity and hTERT transcripts were not expressed in any of the examined cultures and telomeres shortened during the culture period. ALT was not evidenced in the MSCs tested. BM-derived MSCs can be safely expanded in vitro and are not susceptible to malignant transformation, thus rendering these cells suitable for cell therapy approaches.