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Assessing the immunopotency of Toll-like receptor agonists in an in vitro tissue-engineered immunological model

机译:在体外组织工程免疫模型中评估Toll样受体激动剂的免疫力

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摘要

The in vitro Peripheral Tissue Equivalent (PTE) module is a three-dimensional tissue-engineered endothelial cell/collagen matrix culture system, which has been reported to reproduce in vivo physiological conditions and which generates dendritic cells (DC) autonomously. In the present study, we used the PTE module to investigate the immunopotency of Toll-like receptor (TLR) agonists, including polyinosine-polycytidylic acid, Gardiquimod, CpG 2006 and lipopolysaccharide. Application of TLR agonists in the PTE module induced a wide range of cytokines, including interleukins 1α/β, 6, 8 and 10 and tumour necrosis factor-α. Compared with traditional peripheral blood mononuclear cell (PBMC) cultures, the PTE module produced twofold to 100-fold higher levels of cytokine secretion, indicating that it can be a highly sensitive assay system. This increased sensitivity is the result of the natural synergy between the leucocytes and the endothelium. Furthermore, the application of TLR agonists, such as lipopolysaccharide and Gardiquimod, to the PTE module enhanced DC differentiation and promoted DC maturation, as indicated by up-regulated expression of CD83, CD86 and CCR7(CD197). In addition, functional assays indicated PTE-derived DC treated with Gardiquimod, a TLR-7 agonist, significantly augmented anti-tetanus toxoid antibody production. Interestingly, replacing PBMC with purified myeloid cells (CD33+) significantly reduced the responsiveness of the PTE module to TLR stimulation. The reduced sensitivity was partly the result of the removal of plasmacytoid DC that participated in the response to TLR stimulation and sensitization of the PTE module. Overall, the in vitro PTE module clearly demonstrated the effects of TLR agonists on DC generation, maturation and antigen-presenting capacity, and may serve as a sensitive and predictive test bed for the evaluation of adjuvant candidates.
机译:体外外周组织等效(PTE)模块是一个三维组织工程化的内皮细胞/胶原基质培养系统,据报道,该系统可重现体内生理条件并自动生成树突状细胞(DC)。在本研究中,我们使用PTE模块来研究Toll样受体(TLR)激动剂的免疫力,包括多肌苷-聚胞苷酸,加地基莫德,CpG 2006和脂多糖。 TLR激动剂在PTE模块中的应用诱导了广泛的细胞因子,包括白介素1α/β,6、8和10和肿瘤坏死因子-α。与传统的外周血单个核细胞(PBMC)培养相比,PTE模块产生的细胞因子分泌水平高出两倍至100倍,这表明它可以成为高度敏感的测定系统。这种增加的敏感性是白细胞与内皮之间自然协同作用的结果。此外,如CD83,CD86和CCR7(CD197)的表达上调所示,将TLR激动剂(如脂多糖和Gardiquimod)应用于PTE模块可增强DC分化并促进DC成熟。此外,功能测定表明,用TLR-7激动剂Gardiquimod处理的PTE衍生DC大大增加了抗破伤风类毒素抗体的产生。有趣的是,用纯化的髓样细胞(CD33 +)代替PBMC显着降低了PTE模块对TLR刺激的反应性。敏感性降低部分是由于去除了参与TLR刺激和PTE模块增敏反应的浆细胞样DC。总体而言,体外PTE模块清楚地证明了TLR激动剂对DC生成,成熟和抗原呈递能力的影响,并且可以作为评估佐剂候选物的灵敏和预测性试验床。

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