Bone morphogenetic protein 9 ( BMP 9 ) induces effective bone formation from reversibly immortalized multipotent adipose-derived ( iMAD ) mesenchymal stem cells
Bone morphogenetic protein 9 ( BMP 9 ) induces effective bone formation from reversibly immortalized multipotent adipose-derived ( iMAD ) mesenchymal stem cells
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发表时间：
2016
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通讯作者：
Shun Lu;Jing Wang;Jixing Ye;Y. Zou;Yunxiao Zhu;Q. Wei;Xin Wang;Shengli Tang;Hao Liu
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作者：
Shun Lu;Jing Wang;Jixing Ye;Y. Zou;Yunxiao Zhu;Q. Wei;Xin Wang;Shengli Tang;Hao Liu
利用间充质干细胞（MSCs）的再生医学和骨组织工程作为骨和骨骼重建的有效方法具有很大的希望。虽然脂肪组织含有MSC样祖细胞或多能脂肪源性细胞（MAD），但重要的是识别和表征可以有效诱导MAD成骨分化的潜在生物因子。为了克服分离和培养原代MAD的耗时和技术上的挑战性过程，在这里，我们建立和表征可逆永生化的小鼠多能脂肪衍生细胞（iMAD）。分离的小鼠原代腹股沟MAD细胞通过逆转录病毒介导的侧接FRT位点的SV 40 T抗原的表达可逆地永生化。显示iMAD表达最常见的MSC标志物。FLP介导的SV 40 T抗原的去除有效地降低了iMAD的增殖活性和细胞存活，表明永生化是可逆的。使用高度成骨的BMP 9，我们发现iMAD对BMP 9刺激高度响应，表达多种谱系调节剂，并且在BMP 9刺激后在体外经历成骨分化。此外，我们证明BMP-9刺激的iMAD与温度响应性生物可降解支架材料形成坚固的异位骨。总的来说，我们的研究结果表明，可逆永生化的iMAD表现出多能MSC的特征，并且对BMP 9诱导的成骨分化高度响应。因此，iMAD应该为MAD生物学的研究提供有价值的资源，这将最终使我们能够为基于MAD的骨组织工程开发新的和有效的策略。
Regenerative medicine and bone tissue engineering using mesenchymal stem cells (MSCs) hold great promise as an effective approach to bone and skeletal reconstruction. While adipose tissue harbors MSC-like progenitors, or multipotent adipose-derived cells (MADs), it is important to identify and characterize potential biological factors that can effectively induce osteogenic differentiation of MADs. To overcome the time-consuming and technically challenging process of isolating and culturing primary MADs, here we establish and characterize the reversibly immortalized mouse multipotent adipose-derived cells (iMADs). The isolated mouse primary inguinal MAD cells are reversibly immortalized via the retrovirus-mediated expression of SV40 T antigen flanked with FRT sites. The iMADs are shown to express most common MSC markers. FLP-mediated removal of SV40 T antigen effectively reduces the proliferative activity and cell survival of iMADs, indicating the immortalization is reversible. Using the highly osteogenic BMP9, we find that the iMADs are highly responsive to BMP9 stimulation, express multiple lineage regulators, and undergo osteogenic differentiation in vitro upon BMP9 stimulation. Furthermore, we demonstrate that BMP9stimulated iMADs form robust ectopic bone with a thermoresponsive biodegradable scaffold material. Collectively, our results demonstrate that the reversibly immortalized iMADs exhibit the characteristics of multipotent MSCs and are highly responsive to BMP9-induced osteogenic differentiation. Thus, the iMADs should provide a valuable resource for the study of MAD biology, which would ultimately enable us to develop novel and efficacious strategies for MAD-based bone tissue engineering.