Production of CETD transgenic mouse line allowing ablation of any type of specific cell population
Production of CETD transgenic mouse line allowing ablation of any type of specific cell population
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DOI：
10.1002/mrd.20323
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发表时间：
2005-09-01
影响因子：
2.5
通讯作者：
Tanigawa, M
中科院分区：
文献类型：
--
作者：
Sato, M;Tanigawa, M
白喉毒素A链(DT-A)是一种有效的蛋白质合成抑制因子。即使只有一个分子的DT-A也能导致细胞死亡。由组织特异性启动子驱动的DT-A基因被用来实现特定细胞系的遗传消融。然而，这种转基因方法往往会导致无关细胞的异常耗尽。为了避免这种情况，我们建立了一种通过Cre-IoxP系统控制DT-A基因表达来特异性耗尽细胞群体的方法。我们获得了5只携带含有IoxP侧翼增强型绿色荧光蛋白(EGFP)基因和DT-A基因的CETD的转基因小鼠。不出所料，将Cre表达载体导入CETD转基因胎儿的原代培养细胞，可导致大量细胞丢失。由髓鞘碱性蛋白(MBP)启动子控制Cre表达的CETD转基因小鼠与MNCE转基因小鼠杂交获得的双基因(双转基因)小鼠表现出胚胎致死性，提示Cre在胚胎阶段表达。向CETD小鼠静脉注射Cre表达载体可导致肾小球病变，可能是由于肾小球上皮细胞的主要耗尽所致。这种基于Cre-IoxP的细胞消融技术是一种强大而方便的方法，可以产生缺乏任何选定细胞群的小鼠。
Diphtheria, toxin A-chain (DT-A) is a potent inhibitor of protein synthesis. As little as a single molecule of DT-A can result in cell death. DT-A gene driven by a tissue-specific promoter is used to achieve genetic ablation of a particular cell lineage. However, this transgenic approach often results in aberrant depletion of unrelated cells. To avoid this, we established a method for specific depletion of a cell population by controlled expression of the DT-A gene via the Cre-IoxP system. We produced five transgenic mice carrying CETD construct containing IoxP-flanked enhanced green fluorescent protein (EGFP) cDNA and the DT-A gene. Transfection of primary cultured cells derived from CETD transgenic fetus with Cre expression plasmid resulted in extensive cell loss, as expected. Bigenic (double transgenic) offspring obtained by crossbreeding between CETD and MNCE transgenic mice in which Cre expression is controlled by the myelin basic protein (MBP) promoter exhibited embryonic lethality, suggesting expression of Cre at embryonic stages. Intravenous injection of Cre expression vector to CETD mice led to generation of glomerular lesions, probably due to predominant depletion of glomerular epithelial cells. This Cre-IoxP-based cell ablation technology is powerful and convenient method of generating mice lacking any chosen cell population.