Stable Genetic Alterations of b- Catenin and ROR2 Regulate the Wnt Pathway, Affect the Fate of MSCs
Stable Genetic Alterations of b- Catenin and ROR2 Regulate the Wnt Pathway, Affect the Fate of MSCs
复制标题
DOI：
10.1002/jcp.24500
复制
发表时间：
2014-06-01
影响因子：
5.6
通讯作者：
Qiu, Hai-Bo
中科院分区：
文献类型：
--
作者：
Cai, Shi-Xia;Liu, Ai-Ran;Qiu, Hai-Bo
Wnt 通路已被证明对体外间充质干细胞 (MSC) 的命运至关重要，但由于其信号传导缺乏稳定的改变，其在体内 MSC 中的作用仍然知之甚少。在本研究中，我们利用慢病毒载体构建了具有激活和失活的β-连环蛋白（经典Wnt信号通路的关键分子）或ROR2（非经典Wnt5a/ROR2信号通路的关键分子）修饰的长期稳定的mMSCs系。我们发现慢病毒载体介导的转导效率为92.61-97.04%，并且在mMSCs传代20次以上仍保持不变。慢病毒载体转染不仅调节β-catenin或ROR2的mRNA和蛋白表达，还调节核β-catenin积累或分别属于经典Wnt和非经典Wnt5a/ROR2途径的Wnt5a/JNK和Wnt5a/PKC途径。 -Catenin或ROR2基因过表达促进mMSC增殖、迁移和分化为成骨细胞，同时抑制mMSC的成脂分化。相反，β-连环蛋白或 ROR2 基因的失活会产生相反的效果。因此，这些结果证实慢病毒载体转导可以促进mMSCs中Wnt通路的持续有效的基因修饰。本研究提供了一种研究 Wnt 通路对体内 mMSC 命运的影响的方法，并为进一步改进基于 MSC 的疗法提供了方法。 J.细胞。生理学。 229：791-800，2014 年。(c) 2013 年 Wiley 期刊公司。
The Wnt pathways have been shown to be critical for the fate of mesenchymal stem cells (MSCs) in vitro, but their roles in MSCs in vivo remain poorly characterized due to the lack of stable alterations in their signaling. In the present study, we constructed long-term and stable mMSCs lines with activated and inactivated -catenin (the key molecule of the canonical Wnt signaling pathway) or ROR2 (the key molecule of the noncanonical Wnt5a/ROR2 signaling pathway) modifications with lentiviral vectors. We found that the transduction efficiencies mediated by the lentiviral vectors were 92.61-97.04% and were maintained over 20 passages of mMSCs. Transfection by lentiviral vectors not only regulated the mRNA and protein expression of -catenin or ROR2 but also regulated nuclear -catenin accumulation or the Wnt5a/JNK and Wnt5a/PKC pathways belonging to the canonical Wnt and noncanonical Wnt5a/ROR2 pathways, respectively. -Catenin or ROR2 gene overexpression promoted mMSC proliferation, migration and differentiation into osteoblasts, while inhibiting the adipogenic differentiation of mMSCs. In contrast, inactivation of the -catenin or ROR2 genes resulted in the opposite effects. Therefore, these results confirm that lentiviral vector transduction can facilitate sustained and efficient gene modification of the Wnt pathway in mMSCs. This study provides a method to investigate the effects of the Wnt pathway on the fate of mMSCs in vivo and for the further improvement of MSC-based therapies. J. Cell. Physiol. 229: 791-800, 2014. (c) 2013 Wiley Periodicals, Inc.