Kim Hyun-Joo, Cha Gil Sun, Joo Ji-Young, Lee Juyoun, Kim Sung-Jo, Lee Jeongae, Park So Youn, Choi Jeomil
Department of Periodontology, Dental Research Institute, Pusan National University Dental Hospital, Pusan National University School of Dentistry, Yangsan, Korea.
Molecular Recognition Research Center, Korea Institute of Science and Technology, Seoul, Korea.
J Periodontal Implant Sci. 2017 Oct;47(5):292-311. doi: 10.5051/jpis.2017.47.5.292. Epub 2017 Oct 30.
Beyond the limited scope of non-specific polyclonal regulatory T cell (Treg)-based immunotherapy, which depends largely on serendipity, the present study explored a target Treg subset appropriate for the delivery of a novel epitope spreader Pep19 antigen as part of a sophisticated form of immunotherapy with defined antigen specificity that induces immune tolerance.
Human polyclonal CD4CD25CD127 Tregs (127-Tregs) and naïve CD4CD25CD45RA Tregs (45RA-Tregs) were isolated and were stimulated with target peptide 19 (Pep19)-pulsed dendritic cells in a tolerogenic milieu followed by expansion. Low-dose interleukin-2 (IL-2) and rapamycin were added to selectively exclude the outgrowth of contaminating effector T cells (Teffs). The following parameters were investigated in the expanded antigen-specific Tregs: the distinct expression of the immunosuppressive Treg marker Foxp3, epigenetic stability (demethylation in the Treg-specific demethylated region), the suppression of Teffs, expression of the homing receptors CD62L/CCR7, and CD95L-mediated apoptosis. The expanded Tregs were adoptively transferred into an NOD/scid/IL-2Rγ mouse model of collagen-induced arthritis.
Epitope-spreader Pep19 targeting by 45RA-Tregs led to an outstanding suppressive T cell fate characterized by robust expansion, the salient expression of Foxp3, high epigenetic stability, enhanced T cell suppression, modest expression of CD62L/CCR7, and higher resistance to CD95L-mediated apoptosis. After adoptive transfer, the distinct fate of these T cells demonstrated a potent immunotherapeutic capability, as indicated by the complete elimination of footpad swelling, prolonged survival, minimal histopathological changes, and preferential localization of CD4CD25 Tregs at the articular joints in a mechanistic and orchestrated way.
We propose human naïve CD4CD25CD45RA Tregs and the epitope spreader Pep19 as cellular and molecular targets for a novel antigen-specific Treg-based vaccination against collagen-induced arthritis.
非特异性多克隆调节性T细胞（Treg）免疫疗法的作用范围有限，很大程度上依赖于偶然性，本研究探索了一种适合递送新型表位扩展肽Pep19抗原的靶向Treg亚群，作为一种具有明确抗原特异性且能诱导免疫耐受的复杂免疫疗法的一部分。
分离人多克隆CD4⁺CD25⁻CD127⁺ Tregs（127-Tregs）和初始CD4⁺CD25⁻CD45RA⁺ Tregs（45RA-Tregs），在致耐受性环境中用靶向肽19（Pep19）脉冲刺激的树突状细胞刺激，随后进行扩增。添加低剂量白细胞介素-2（IL-2）和雷帕霉素以选择性排除污染的效应T细胞（Teffs）的生长。在扩增的抗原特异性Tregs中研究以下参数：免疫抑制性Treg标志物Foxp3的独特表达、表观遗传稳定性（Treg特异性去甲基化区域的去甲基化）、对Teffs的抑制、归巢受体CD62L/CCR7的表达以及CD95L介导的细胞凋亡。将扩增的Tregs过继转移到胶原诱导性关节炎的NOD/scid/IL-2Rγ小鼠模型中。
45RA-Tregs对表位扩展肽Pep19的靶向作用导致显著的抑制性T细胞命运，其特征为强劲扩增、Foxp3的显著表达、高表观遗传稳定性、增强的T细胞抑制、适度的CD62L/CCR7表达以及对CD95L介导的细胞凋亡的更高抗性。过继转移后，这些T细胞的独特命运显示出强大的免疫治疗能力，表现为足垫肿胀完全消除、生存期延长、组织病理学变化最小以及CD4⁺CD25⁺ Tregs以机械协调的方式优先定位于关节。
我们提出人初始CD4⁺CD25⁻CD45RA⁺ Tregs和表位扩展肽Pep19作为针对胶原诱导性关节炎的新型基于抗原特异性Treg的疫苗接种的细胞和分子靶点。
