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Tetramer-Based Enrichment of Preexisting Anti-AAV8 CD8+ T Cells in Human Donors Allows the Detection of a TEMRA Subpopulation.
Frontiers in Immunology ( IF 5.7 ) Pub Date : 2020-01-21 , DOI: 10.3389/fimmu.2019.03110
Céline Vandamme 1 , Rebecca Xicluna 1 , Leslie Hesnard 2 , Marie Devaux 1 , Nicolas Jaulin 1 , Mickaël Guilbaud 1 , Johanne Le Duff 1 , Célia Couzinié 1 , Philippe Moullier 1 , Xavier Saulquin 2 , Oumeya Adjali 1
Affiliation  

Pre-existing immunity to AAV capsid may compromise the safety and efficiency of rAAV-mediated gene transfer in patients. Anti-capsid cytotoxic immune responses have proven to be a challenge to characterize because of the scarcity of circulating AAV-specific CD8+ T lymphocytes which can seldom be detected with conventional flow cytometry or ELISpot assays. Here, we used fluorescent MHC class I tetramers combined with magnetic enrichment to detect and phenotype AAV8-specific CD8+ T cells in human PBMCs without prior amplification. We showed that all healthy individuals tested carried a pool of AAV8-specific CD8+ T cells with a CD45RA+ CCR7- terminally-differentiated effector memory cell (TEMRA) fraction. Ex vivo frequencies of total AAV-specific CD8+ T cells were not predictive of IFNγ ELISpot responses but interestingly we evidenced a correlation between the proportion of TEMRA cells and IFNγ ELISpot positive responses. TEMRA cells may then play a role in recombinant AAV-mediated cytotoxicity in patients with preexisting immunity. Overall, our results encourage the development of new methods combining increased detection sensitivity of AAV-specific T cells and their poly-functional assessment to better characterize and monitor AAV capsid-specific cellular immune responses in the perspective of rAAV-mediated clinical trials.

中文翻译:

基于四聚体的人类供体中现有抗AAV8 CD8 + T细胞的富集允许检测TEMRA亚群。

预先存在的对AAV衣壳的免疫力可能会危及rAAV介导的基因转移患者的安全性和效率。由于缺乏循环AAV特异性CD8 + T淋巴细胞的稀缺性,抗衣壳细胞毒性免疫反应已被证明是表征的挑战,而传统的流式细胞仪或ELISpot分析很少能检测到这种抗体。在这里,我们使用荧光MHC I类四聚体与磁富集相结合来检测人PBMC中的AAV8特异性CD8 + T细胞并表型,而无需事先扩增。我们显示,所有接受测试的健康个体均携带AAV8特异性CD8 + T细胞库,并带有CD45RA + CCR7-末端分化的效应记忆细胞(TEMRA)级分。总的AAV特异性CD8 + T细胞的离体频率不能预测IFNγELISpot反应,但有趣的是,我们证明了TEMRA细胞比例与IFNγELISpot阳性反应之间存在相关性。然后,TEMRA细胞可能在具有先前免疫功能的患者中在重组AAV介导的细胞毒性中发挥作用。总体而言,我们的结果鼓励开发新方法,结合rAAV介导的临床试验的观点,结合提高AAV特异性T细胞的检测灵敏度及其多功能评估,以更好地表征和监测AAV衣壳特异性细胞免疫反应。
更新日期:2020-01-22
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