Isolation of murine bone marrow derived mesenchymal stem cells using Twist2 Cre transgenic mice.
Isolation of murine bone marrow derived mesenchymal stem cells using Twist2 Cre transgenic mice.
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DOI：
10.1016/j.bone.2010.07.022
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发表时间：
2010-11
期刊：
影响因子：
4.1
通讯作者：
Maye, Peter
中科院分区：
文献类型：
--
作者：
Liu, Yaling;Wang, Liping;Fatahi, Reza;Kronenberg, Mark;Kalajzic, Ivo;Rowe, David;Li, Yingcui;Maye, Peter
虽然人骨髓源性间充质干细胞（BMSC）因其潜在的治疗价值而引起极大的兴趣，但其小鼠等效物仍然是一个重要的基础研究模型，可以为这种祖细胞群体的生物学提供重要的见解。在这里，我们提出了一种新的转基因策略，允许在基质细胞培养的早期阶段选择性地识别和分离小鼠BMSCs。该策略涉及将Twist 2-Cre小鼠与Cre报告小鼠如Z/EG或Ai 9杂交，其在Cre介导的终止序列切除后分别表达EGFP或Tomato荧光蛋白。使用这种方法，我们鉴定了在集落形成的最早阶段存在的粘附荧光蛋白+细胞群体（T2 C+），并且在培养的第5天代表总细胞群体的~20%。通过流式细胞术的细胞表面分析显示，T2 C+细胞对SCA 1和CD 29呈高度阳性，对CD 45、CD 117、TIE 2和TER 119呈阴性。通过FACS分离T2 C+细胞，选择具有骨骼潜能的细胞群，其可被引导分化成成骨细胞、脂肪细胞或软骨细胞。我们还在颅骨骨缺损模型中证明，T2 C+细胞保留了成骨修复的强大功效，并可以支持造血环境。总的来说，这些研究提供了证据表明，Twist 2-Cre x Cre报告基因育种策略可用于阳性鉴定和分离多能鼠BMSC。
While human bone marrow derived mesenchymal stem cells (BMSCs) are of great interest for their potential therapeutic value, its murine equivalent remains an important basic research model that can provide critical insights into the biology of this progenitor cell population. Here we present a novel transgenic strategy that allowed for the selective identification and isolation of murine BMSCs at the early stages of stromal cell culture. This strategy involved crossing Twist2 –Cre mice with Cre reporter mice such as Z/EG or Ai9, which express EGFP or Tomato fluorescent protein, respectively, upon Cre mediated excision of a stop sequence. Using this approach, we identified an adherent fluorescent protein+ cell population (T2C+) that is present during the earliest stages of colony formation and by day 5 of culture represents ~20% of the total cell population. Cell surface profiling by flow cytometry showed that T2C+ cells are highly positive for SCA1 and CD29 and negative for CD45, CD117, TIE2, and TER119. Isolation of T2C+ cells by FACS selected for a cell population with skeletal potential that can be directed to differentiate into osteoblasts, adipocytes, or chondrocytes. We also demonstrated in a calvarial bone defect model that T2C+ cells retain a strong efficacy for osteogenic repair and can support a hematopoietic environment. Collectively, these studies provide evidence that the Twist2-Cre x Cre reporter breeding strategy can be used to positively identify and isolate multipotent murine BMSCs.
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影响因子：
64.5
作者：
Yang, J;Mani, SA;Weinberg, RA
通讯作者：
Weinberg, RA
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64.5
作者：
Sacchetti, Benedetto;Funari, Alessia;Bianco, Paolo
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20.3
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GRONTHOS, S;GRAVES, SE;SIMMONS, PJ
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SIMMONS, PJ
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11.8
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通讯作者：
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