Dipartimento di Biologia Animale, Laboratorio di Biologia dello Sviluppo, Universita' degli Studi di Pavia, Piazza Botta 9, 27100, Pavia, Italy.
Cytotechnology. 2008 Sep;58(1):17-23. doi: 10.1007/s10616-008-9164-x. Epub 2008 Sep 30.
Although mouse embryonic stem cell lines (mESCs) have been established since 1981, systematic studies about chromosomal changes during culture are lacking. In this study, we report the results of a cytogenetic analysis performed on three mESC lines (named UPV02, UPV06 and UPV08) cultured for a period of 3 months. At time intervals, the variation of the chromosome number together with the expression of markers of the undifferentiated status, i.e., OCT-4, SSEA-1, FOM-1 and alkaline phosphatase activity, were determined. The three mESC lines showed a progressive loss of euploid metaphases during the 3 months period of culture. Chromosome abnormalities were accumulated at the latest passages analysed. Metacentric chromosomes were the most frequent chromosome abnormality observed throughout the period of culture. Interestingly, in coincidence with, or few passages after, the drop of euploidy, the alkaline phosphatase activity was partially or totally lost, whereas the OCT-4, SSEA-1 and FOM-1 stem markers were always positive throughout the period of culture. Our results remark the necessity to perform the karyotype analysis during culture in order to develop new culture conditions to maintain the correct chromosome complement in long-term culture of mESC lines.
尽管小鼠胚胎干细胞系 (mESCs) 自 1981 年以来已经建立，但在培养过程中关于染色体变化的系统研究还很缺乏。在这项研究中，我们报告了对三个培养了 3 个月的 mESC 系 (命名为 UPV02、UPV06 和 UPV08) 进行细胞遗传学分析的结果。在不同时间间隔，我们测定了染色体数目的变化以及未分化状态标志物 OCT-4、SSEA-1、FOM-1 和碱性磷酸酶活性的表达。在培养的 3 个月期间，这三个 mESC 系显示出逐渐丢失整倍体中期的趋势。在分析的最新传代中积累了染色体异常。在整个培养期间，着丝粒染色体是最常见的染色体异常。有趣的是，与整倍体丢失的时间一致，或在其之后的几个传代中，碱性磷酸酶活性部分或完全丢失，而 OCT-4、SSEA-1 和 FOM-1 干细胞标志物在整个培养期间始终为阳性。我们的结果表明，为了开发新的培养条件以维持 mESC 系的长期培养中正确的染色体组成，有必要在培养过程中进行核型分析。
