Manitoba Centre for Proteomics and Systems Biology, Department of Internal Medicine, University of Manitoba, 799 John Buhler Research Centre, 715 McDermot Avenue, Winnipeg, MB, R3E3P4, Canada.
Arthritis Res Ther. 2011 Aug 11;13(4):R129. doi: 10.1186/ar3440.
Innate defence regulator (IDR) peptides are synthetic cationic peptides, variants of naturally occurring innate immune effector molecules known as host defence peptides. IDR peptides were recently demonstrated to limit infection-associated inflammation selectively without compromising host innate immune functions. This study examined the impact of a 12-amino acid IDR peptide, IDR-1002, in pro-inflammatory cytokine interleukin (IL)-1β-induced responses in synovial fibroblasts, a critical cell type in the pathogenesis of inflammatory arthritis.
Human fibroblast-like synoviocytes (FLS) were stimulated with IL-1β in the presence and absence of IDR-1002. Production of enzyme matrix metalloproteinase-3 (MMP-3) and IL-1-receptor antagonist (IL-1RA) was monitored by enzyme-linked immunosorbent assay (ELISA), and various chemokines were evaluated by using multiplex cytometric bead array. Transcriptional responses were analyzed by quantitative real-time PCR. The impact on IL-1β-induced proteome was investigated by quantitative proteomics by using isobaric tags. IL-1β-induced pathways altered by IDR-1002 implicated by the proteomics analyses were further investigated by using various immunochemical assays. Cellular uptake of the peptide was monitored by using a biotinylated IDR-1002 peptide followed by microscopy probing with streptavidin-Alexa Fluor.
This study demonstrated that IDR-1002 suppressed the production of IL-1β-induced MMP-3 and monocyte chemotactic protein-1 (MCP-1); in contrast, IDR-1002 enhanced the production of IL-1RA, without neutralizing all chemokine responses. IDR-1002 altered the IL-1β-induced proteome primarily by altering the expression of members of nuclear factor kappa-B (NF-κB) and c-Jun N-terminal kinase (JNK) pathways. The proteomics data also suggested that IDR-1002 was altering the transcription factor HNF-4α-mediated responses, known to be critical in metabolic regulation. With various immunochemical assays, it was further demonstrated that IL-1β-induced NF-κB, JNK, and p38 mitogen-activated protein kinase (MAPK) activations were significantly suppressed by IDR-1002.
This study demonstrates the ability of an innate immune-modulatory IDR-peptide to influence the IL-1β-induced regulatory pathways and selectively to suppress inflammatory responses in synovial fibroblasts. The results of this study provide a rationale for examining the use of IDR-peptides as potential therapeutic candidates for chronic inflammatory diseases such as inflammatory arthritis.
先天防御调节剂（IDR）肽是合成的阳离子肽，是天然存在的先天免疫效应分子的变体，称为宿主防御肽。最近的研究表明，IDR 肽可以选择性地限制感染相关的炎症，而不损害宿主的先天免疫功能。本研究检测了一种 12 个氨基酸的 IDR 肽 IDR-1002 在白细胞介素（IL）-1β诱导的滑膜成纤维细胞（炎症性关节炎发病机制中的关键细胞类型）中的促炎细胞因子反应中的作用。
用 IL-1β刺激人成纤维样滑膜细胞（FLS），并在存在和不存在 IDR-1002 的情况下刺激。通过酶联免疫吸附试验（ELISA）监测酶基质金属蛋白酶-3（MMP-3）和白细胞介素-1 受体拮抗剂（IL-1RA）的产生，并用多重流式细胞术测定各种趋化因子。通过定量实时 PCR 分析转录反应。通过使用等压标签进行定量蛋白质组学研究，研究 IDR-1002 对 IL-1β诱导的蛋白质组的影响。通过各种免疫化学测定进一步研究蛋白质组学分析暗示的 IDR-1002 改变的 IL-1β诱导途径。通过使用生物素化 IDR-1002 肽并随后用链霉亲和素-Alexa Fluor 进行显微镜探测，监测肽的细胞摄取。
本研究表明，IDR-1002 抑制了 IL-1β诱导的 MMP-3 和单核细胞趋化蛋白-1（MCP-1）的产生；相反，IDR-1002 增强了 IL-1RA 的产生，而不中和所有趋化因子的反应。IDR-1002 主要通过改变核因子 kappa-B（NF-κB）和 c-Jun N 端激酶（JNK）途径的成员的表达来改变 IL-1β 诱导的蛋白质组。蛋白质组学数据还表明，IDR-1002 改变了转录因子 HNF-4α 介导的反应，这在代谢调节中是至关重要的。通过各种免疫化学测定，进一步证明 IDR-1002 显著抑制了 IL-1β 诱导的 NF-κB、JNK 和 p38 丝裂原活化蛋白激酶（MAPK）的激活。
本研究证明了一种先天免疫调节 IDR-肽能够影响 IL-1β 诱导的调节途径，并选择性地抑制滑膜成纤维细胞的炎症反应。本研究的结果为研究 IDR-肽作为炎症性关节炎等慢性炎症性疾病的潜在治疗候选物提供了依据。
