Havenar-Daughton Colin, Reiss Samantha M, Carnathan Diane G, Wu Jennifer E, Kendric Kayla, Torrents de la Peña Alba, Kasturi Sudhir Pai, Dan Jennifer M, Bothwell Marcella, Sanders Rogier W, Pulendran Bali, Silvestri Guido, Crotty Shane
Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037; Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery, La Jolla, CA 92037;
Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037;
J Immunol. 2016 Aug 1;197(3):994-1002. doi: 10.4049/jimmunol.1600320. Epub 2016 Jun 22.
A range of current candidate AIDS vaccine regimens are focused on generating protective HIV-neutralizing Ab responses. Many of these efforts rely on the rhesus macaque animal model. Understanding how protective Ab responses develop and how to increase their efficacy are both major knowledge gaps. Germinal centers (GCs) are the engines of Ab affinity maturation. GC T follicular helper (Tfh) CD4 T cells are required for GCs. Studying vaccine-specific GC Tfh cells after protein immunizations has been challenging, as Ag-specific GC Tfh cells are difficult to identify by conventional intracellular cytokine staining. Cytokine production by GC Tfh cells may be intrinsically limited in comparison with other Th effector cells, as the biological role of a GC Tfh cell is to provide help to individual B cells within the GC, rather than secreting large amounts of cytokines bathing a tissue. To test this idea, we developed a cytokine-independent method to identify Ag-specific GC Tfh cells. RNA sequencing was performed using TCR-stimulated GC Tfh cells to identify candidate markers. Validation experiments determined CD25 (IL-2Rα) and OX40 to be highly upregulated activation-induced markers (AIM) on the surface of GC Tfh cells after stimulation. In comparison with intracellular cytokine staining, the AIM assay identified >10-fold more Ag-specific GC Tfh cells in HIV Env protein-immunized macaques (BG505 SOSIP). CD4 T cells in blood were also studied. In summary, AIM demonstrates that Ag-specific GC Tfh cells are intrinsically stingy producers of cytokines, which is likely an essential part of their biological function.
一系列当前的候选艾滋病疫苗方案都聚焦于产生具有保护性的HIV中和抗体反应。其中许多努力都依赖于恒河猴动物模型。了解保护性抗体反应如何产生以及如何提高其效力都是主要的知识空白。生发中心（GCs）是抗体亲和力成熟的引擎。GCs需要T滤泡辅助（Tfh）CD4 T细胞。在蛋白质免疫后研究疫苗特异性GC Tfh细胞一直具有挑战性，因为抗原特异性GC Tfh细胞难以通过传统的细胞内细胞因子染色来识别。与其他Th效应细胞相比，GC Tfh细胞产生细胞因子的能力可能内在受限，因为GC Tfh细胞的生物学作用是为GC内的单个B细胞提供帮助，而不是分泌大量细胞因子浸润组织。为了验证这一观点，我们开发了一种不依赖细胞因子的方法来识别抗原特异性GC Tfh细胞。使用TCR刺激的GC Tfh细胞进行RNA测序以识别候选标志物。验证实验确定CD25（IL-2Rα）和OX40是刺激后GC Tfh细胞表面高度上调的激活诱导标志物（AIM）。与细胞内细胞因子染色相比，AIM分析在HIV Env蛋白免疫的猕猴（BG505 SOSIP）中识别出的抗原特异性GC Tfh细胞多10倍以上。还研究了血液中的CD4 T细胞。总之，AIM表明抗原特异性GC Tfh细胞本质上是细胞因子的少量生产者，这可能是其生物学功能的重要组成部分。
