Department of Chemical Engineering, University of Michigan, Ann Arbor, Michigan, USA.
Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan, USA.
Biotechnol Bioeng. 2023 Jan;120(1):284-296. doi: 10.1002/bit.28252. Epub 2022 Oct 21.
Immune-mediated hypersensitivities such as autoimmunity, allergy, and allogeneic graft rejection are treated with therapeutics that suppress the immune system, and the lack of specificity is associated with significant side effects. The delivery of disease-relevant antigens (Ags) by carrier systems such as poly(lactide-co-glycolide) nanoparticles (PLG-Ag) and carbodiimide (ECDI)-fixed splenocytes (SP-Ag) has demonstrated Ag-specific tolerance induction in model systems of these diseases. Despite therapeutic outcomes by both platforms, tolerance is conferred with different efficacy. This investigation evaluated Ag loading and total particle dose of PLG-Ag on Ag presentation in a coculture system of dendritic cells (DCs) and Ag-restricted T cells, with SP-Ag employed as a control. CD25 expression was observed in nearly all T cells even at low concentrations of PLG-Ag, indicating efficient presentation of Ag by dendritic cells. However, the secretion of IL-2, Th1, and Th2 cytokines (IFNγ and IL-4, respectively) varied depending on PLG-Ag concentration and Ag loading. Concentration escalation of soluble Ag resulted in an increase in IL-2 and IFNγ and a decrease in IL-4. Treatment with PLG-Ag followed a similar trend but with lower levels of IL-2 and IFNγ secreted. Transcriptional Activity CEll ARrays (TRACER) were employed to measure the real-time transcription factor (TF) activity in Ag-presenting DCs. The kinetics and magnitude of TF activity was dependent on the Ag delivery method, concentration, and Ag loading. Ag positively regulated IRF1 activity and, as carriers, NPs and ECDI-treated SP negatively regulated this signaling. The effect of Ag loading and dose on tolerance induction were corroborated in vivo using the delayed-type hypersensitivity (DTH) and experimental autoimmune encephalomyelitis (EAE) mouse models where a threshold of 8 μg/mg Ag loading and 0.5 mg PLG-Ag dose were required for tolerance. Together, the effect of Ag loading and dosing on in vitro and in vivo immune regulation provide useful insights for translating Ag-carrier systems for the clinical treatment of immune disorders.
免疫介导的超敏反应，如自身免疫、过敏和同种异体移植物排斥，可通过抑制免疫系统的治疗方法进行治疗，但其缺乏特异性会导致严重的副作用。将疾病相关抗原 (Ag) 通过载体系统（如聚（乳酸-共-乙醇酸）纳米颗粒 (PLG-Ag) 和碳二亚胺 (ECDI) 固定的脾细胞 (SP-Ag)）递送给机体，在这些疾病的模型系统中已证明可诱导 Ag 特异性耐受。尽管这两种平台都取得了治疗效果，但它们诱导的耐受效果却有所不同。本研究评估了 PLG-Ag 的 Ag 加载量和总颗粒剂量对树突状细胞 (DC) 和 Ag 限制的 T 细胞共培养系统中 Ag 呈递的影响，并用 SP-Ag 作为对照。在低浓度的 PLG-Ag 下，几乎所有 T 细胞都观察到 CD25 的表达，这表明 DC 有效呈递了 Ag。然而，IL-2、Th1 和 Th2 细胞因子（分别为 IFNγ 和 IL-4）的分泌因 PLG-Ag 浓度和 Ag 加载量而异。可溶性 Ag 浓度的增加导致 IL-2 和 IFNγ 的增加以及 IL-4 的减少。PLG-Ag 的处理则呈现出相似的趋势，但分泌的 IL-2 和 IFNγ 水平较低。采用转录因子活性细胞分析（TRACER）来测量 Ag 呈递 DC 中的实时转录因子 (TF) 活性。TF 活性的动力学和幅度取决于 Ag 递呈方式、浓度和 Ag 加载量。Ag 正向调节 IRF1 活性，而作为载体的 NPs 和 ECDI 处理的 SP 则负向调节该信号。通过迟发型超敏反应 (DTH) 和实验性自身免疫性脑脊髓炎 (EAE) 小鼠模型在体内验证了 Ag 加载量和剂量对诱导耐受的影响，其中需要 8μg/mg Ag 加载量和 0.5mg PLG-Ag 剂量作为诱导耐受的阈值。综上所述，Ag 加载量和剂量对体外和体内免疫调节的影响为将 Ag-载体系统转化为免疫疾病的临床治疗提供了有用的见解。
