Selective modification of antigen-specific T cells by RNA electroporation
Selective modification of antigen-specific T cells by RNA electroporation
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DOI：
10.1089/hum.2007.115
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发表时间：
2008-05-01
影响因子：
4.2
通讯作者：
Sampson, John H.
中科院分区：
文献类型：
--
作者：
Mitchell, Duane A.;Karikari, Isaac;Sampson, John H.
已经观察到，通过RNA电穿孔有效转染T细胞需要用促分裂原或通过抗CD 3抗体刺激预先活化T细胞。我们假设，T细胞活化的这一要求可以被利用来表达对抗原脉冲树突状细胞应答的活化T细胞内的标记基因，并允许选择性富集和修饰抗原特异性T细胞。使用编码绿色荧光蛋白的mRNA的电穿孔作为标记基因，我们证明RNA电穿孔可以有效地允许从响应于巨细胞病毒pp 65抗原脉冲的树突状细胞的大量培养物中分离巨细胞病毒特异性CD 8（+）和CD 4（+）T细胞。此外，我们证明巨细胞病毒特异性T细胞可以通过C-X-C趋化因子受体CXCR 2的RNA转染进行功能修饰，以在体外和体内有效地向各种CXCR 2特异性趋化因子迁移。这些研究证明了RNA转染作为纯化和选择性修饰抗原特异性T细胞的功能以用于过继免疫治疗的简单方法的实用性，并且重要地提供了通过RNA转染瞬时表达蛋白质是调节活化T细胞的体内功能的有效手段的证据。
It has been observed that the efficient transfection of T cells by RNA electroporation requires prior activation of T cells with mitogens or by anti-CD3 antibody stimulation. We hypothesized that this requirement for T cell activation could be leveraged to express marker genes within activated T cells responding to antigen-pulsed dendritic cells and allow for the selective enrichment and modification of antigen-specific T cells. Using electroporation of mRNA encoding green fluorescent protein as a marker gene, we demonstrate that RNA electroporation can efficiently allow for the separation of cytomegalovirus-specific CD8(+) and CD4(+) T cells from bulk culture responding to cytomegalovirus pp65 antigen-pulsed dendritic cells. Furthermore, we demonstrate that cytomegalovirus-specific T cells can be functionally modified by RNA transfection of the C-X-C chemokine receptor, CXCR2, to migrate efficiently toward a variety of CXCR2-specific chemokines in vitro and in vivo. These studies demonstrate the utility of RNA transfection as a simple method by which to purify and selectively modify the function of antigen-specific T cells for use in adoptive immunotherapy, and importantly provide evidence that transient expression of proteins by RNA transfection is an efficient means of modulating the in vivo function of activated T cells.