Wang B, Levelt C, Salio M, Zheng D, Sancho J, Liu C P, She J, Huang M, Higgins K, Sunshine M J
Division of Immunology, Beth Israel Hospital, Harvard Medical School, Boston, MA 02115, USA.
Int Immunol. 1995 Mar;7(3):435-48. doi: 10.1093/intimm/7.3.435.
We have reported previously that mice carrying > 30 copies of the human CD3 epsilon transgene completely lose their T lymphocytes and NK cells (36). Here we demonstrate by immunohistology that in the most severely immunodeficient mouse, tg epsilon 26, the thymus is very small, has sizeable vacuoles and does not contain recognizable T lymphocytes except for a small percentage of Thy-1+ cells and B cells. Cell surface phenotyping and TCR alpha and -beta rearrangement studies confirm that the arrest in T lymphocyte development precedes the arrest in rag-1null, rag-2null and TCR beta nuli mice. Since the T cell progenitors in which the arrest occurred were absent in the transgenic mice, indirect approaches were taken to examine the causes of the block in T cell development. Analyses of 12 independently established mutant mouse lines, generated with five different transgenic constructs, revealed that the severity of the abrogation in T cell development was dependent on the number of copies of transgenes. Since the number of transgene copies generally correlated with the levels of expression of the transgenic CD3 epsilon proteins, we concluded that over-expression of the CD3 epsilon protein was the likely cause of the block in T lymphocyte development. The T cell immunodeficiency was caused by either the human or the murine CD3 epsilon protein. Since transgene coded mRNAs were found in significantly higher quantities than endogenous CD3 epsilon mRNAs in fetal thymi on days 13 and 14 of gestation, over-expression took place very early in development, probably prematurely. Over-expression of the CD3 epsilon transgene in thymocyte precursors may therefore affect T lymphocyte development in the absence of TCR and possibly in the absence of the other CD3 proteins. More importantly, over-expression of the CD3 epsilon protein in thymocytes of mice with a low copy number of transgenes had a significant effect on late thymic development. Over-expression of the CD3 epsilon protein in immature thymocytes mimicked the effects caused by exposure of CD4- CD8- thymocytes to anti-CD3 epsilon treatment: apoptosis and lack of TCR beta expression. We therefore speculate that in the homozygous tg epsilon 26 animals the arrest in T cell development was caused by excessive signal transduction events rather than by a toxic effect of the transgenic protein.
我们之前报道过，携带超过30个拷贝的人CD3ε转基因的小鼠完全失去了它们的T淋巴细胞和NK细胞(36)。在这里，我们通过免疫组织学证明，在免疫缺陷最严重的小鼠tg epsilon 26中，胸腺非常小，有相当大的空泡，除了一小部分Thy-1+细胞和B细胞外，不含有可识别的T淋巴细胞。细胞表面表型分析以及TCRα和β重排研究证实，T淋巴细胞发育的停滞先于rag-1基因敲除、rag-2基因敲除和TCRβ基因敲除小鼠中的停滞。由于转基因小鼠中不存在发生停滞的T细胞祖细胞，因此采用间接方法来研究T细胞发育受阻的原因。对用五种不同转基因构建体产生的12个独立建立的突变小鼠品系的分析表明，T细胞发育废除的严重程度取决于转基因的拷贝数。由于转基因拷贝数通常与转基因CD3ε蛋白的表达水平相关，我们得出结论，CD3ε蛋白的过度表达可能是T淋巴细胞发育受阻的原因。T细胞免疫缺陷是由人或小鼠的CD3ε蛋白引起的。由于在妊娠第13天和第14天的胎儿胸腺中发现转基因编码的mRNA数量明显高于内源性CD3ε mRNA，过度表达在发育早期就发生了，可能过早。因此，胸腺细胞前体中CD3ε转基因的过度表达可能在没有TCR且可能没有其他CD3蛋白的情况下影响T淋巴细胞的发育。更重要的是，转基因拷贝数低的小鼠胸腺细胞中CD3ε蛋白的过度表达对胸腺后期发育有显著影响。未成熟胸腺细胞中CD3ε蛋白的过度表达模拟了CD4-CD8-胸腺细胞暴露于抗CD3ε处理所引起的效应：细胞凋亡和TCRβ表达缺失。因此，我们推测在纯合tg epsilon 26动物中，T细胞发育的停滞是由过度的信号转导事件引起的，而不是由转基因蛋白的毒性作用引起的。
