Conditionally immortalized mouse embryonic fibroblasts retain proliferative activity without compromising multipotent differentiation potential.
Conditionally immortalized mouse embryonic fibroblasts retain proliferative activity without compromising multipotent differentiation potential.
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条件永生化小鼠胚胎成纤维细胞保留增殖活性而不影响多能分化潜能
DOI：
10.1371/journal.pone.0032428
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
He TC
中科院分区：
文献类型：
--
作者：
Huang E;Bi Y;Jiang W;Luo X;Yang K;Gao JL;Gao Y;Luo Q;Shi Q;Kim SH;Liu X;Li M;Hu N;Liu H;Cui J;Zhang W;Li R;Chen X;Shen J;Kong Y;Zhang J;Wang J;Luo J;He BC;Wang H;Reid RR;Luu HH;Haydon RC;Yang L;He TC
间充质干细胞（Mesenchymal stem cells，MSCs）是一种多能细胞，存在于多种组织中，可分化为骨、软骨和脂肪等多种细胞系。尽管MSC在基础和转化研究中引起了极大的关注，但原代MSC在培养中的寿命有限，这阻碍了MSC的更广泛应用。在这里，我们调查，如果小鼠间充质祖细胞可以有条件地永生化与SV 40大T抗原，并保持长期的细胞增殖，而不损害他们的多能性。利用表达两侧带有Cre/loxP位点的SV 40大T抗原的系统，我们证明了SV 40大T抗原可以有效地永生化小鼠胚胎成纤维细胞（MEFs）。条件永生化的MEFs（iMEFs）表现出增强的增殖活性并维持长期的细胞增殖，这可以被Cre重组酶逆转。iMEFs表达大多数MSC标志物并保留多能性，因为它们可以在体外和体内适当的分化条件下分化成成骨、软骨和脂肪谱系。SV 40大T的去除降低了iMEFs的分化潜力，可能是由于祖细胞扩增减少。此外，iMEF在皮下注射到无胸腺裸鼠中时显然不具有致瘤性。因此，条件永生化的iMEF不仅维持长期细胞增殖，而且保留分化成多个谱系的能力。我们的研究结果表明，使用SV 40大T抗原的可逆永生化策略可能是一种有效和安全的方法，以建立长期的细胞培养的初级间充质祖细胞的基础和转化研究，以及潜在的临床应用。
Mesenchymal stem cells (MSCs) are multipotent cells which reside in many tissues and can give rise to multiple lineages including bone, cartilage and adipose. Although MSCs have attracted significant attention for basic and translational research, primary MSCs have limited life span in culture which hampers MSCs' broader applications. Here, we investigate if mouse mesenchymal progenitors can be conditionally immortalized with SV40 large T antigen and maintain long-term cell proliferation without compromising their multipotency. Using the system which expresses SV40 large T antigen flanked with Cre/loxP sites, we demonstrate that mouse embryonic fibroblasts (MEFs) can be efficiently immortalized by SV40 large T antigen. The conditionally immortalized MEFs (iMEFs) exhibit an enhanced proliferative activity and maintain long-term cell proliferation, which can be reversed by Cre recombinase. The iMEFs express most MSC markers and retain multipotency as they can differentiate into osteogenic, chondrogenic and adipogenic lineages under appropriate differentiation conditionsin vitroandin vivo. The removal of SV40 large T reduces the differentiation potential of iMEFs possibly due to the decreased progenitor expansion. Furthermore, the iMEFs are apparently not tumorigenic when they are subcutaneously injected into athymic nude mice. Thus, the conditionally immortalized iMEFs not only maintain long-term cell proliferation but also retain the ability to differentiate into multiple lineages. Our results suggest that the reversible immortalization strategy using SV40 large T antigen may be an efficient and safe approach to establishing long-term cell culture of primary mesenchymal progenitors for basic and translational research, as well as for potential clinical applications.
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DOI：
10.1073/pnas.0401596101
发表时间：
2004-11-02
影响因子：
11.1
作者：
Coles, BLK;Angénieux, B;van der Kooy, D
通讯作者：
van der Kooy, D
影响因子：
5.2
作者：
Djouad, Farida;Charbonnier, Louis-Marie;Noel, Daniele
通讯作者：
Noel, Daniele
影响因子：
5.6
作者：
AARONSON, SA;TODARO, GJ
通讯作者：
TODARO, GJ
影响因子：
56.9
作者：
He, TC;Sparks, AB;Kinzler, KW
通讯作者：
Kinzler, KW
影响因子：
5.2
作者：
in't Anker, PS;Scherjon, SA;Kanhai, HHH
通讯作者：
Kanhai, HHH