Soluble IL-1RAcP proteins as inhibitors of inflammation

描述（由申请人提供）： 皮肤炎症涉及细胞因子的局部产生，细胞因子介导白细胞的募集以及角质形成细胞和皮肤中其他细胞的基因表达和细胞表型的改变。白细胞介素-1（IL-1）是局部和全身炎症的重要介质。IL-1通过涉及I型IL-1受体（IL-1 RI）和膜结合IL-1受体辅助蛋白（mIL-1 RAcP）的受体复合物起作用。IL-1 RAcP的可溶性同种型（sIL-1 RAcP）似乎是IL-1信号传导的抑制剂。主要研究者已经鉴定了IL-1 RAcP的新的可溶性同种型，sIL-1 RAcP-β，其具有独特的C-末端。最近发现的与IL-1相关的细胞因子似乎具有与IL-1相似的活性，但通过不同的膜受体IL-1受体相关蛋白2（IL-1 Rrp 2）起作用。IL-1 Rrp 2主要在上皮组织中表达，并且初步数据表明不同的IL-1家族成员通过在原代人角质形成细胞中诱导不同的细胞因子表达谱来协调不同的免疫应答。有趣的是，IL-1 Rrp 2还利用mIL-1 RAcP进行细胞内信号传导。主要假设是可溶性IL-1 RAcP的两种不同亚型，sIL-1 RAcP和sIL-1 RAcP-1，|3，可以抑制所有IL-1家族成员或其子集的活性。此外，IL-1 RAcP的三种不同亚型的比例表达可能决定细胞对炎症刺激的反应。由于IL-1 Rrp 2主要在上皮细胞中表达，所涉及的机制可能在皮肤稳态和炎症中起重要作用。将表达和纯化重组可溶性IL-1 RAcP蛋白。将用激动剂IL-1家族成员处理人角质形成细胞，并使用定量逆转录PCR和ELISA测定来检查基因表达的激活。两种可溶性IL-1 RAcP同种型的活性将在与单独细胞因子的共处理实验中检查。在用已知的IL-1 RAcP比例表达调节剂和IL-1家族成员处理后，测定角质形成细胞中三种不同IL-1 RAcP同种型和IL-1家族成员的表达。本研究将为IL-1相关细胞因子和两种可溶性IL-1 RacP亚型的功能提供新的信息。最终目标是阐明调节炎症的分子机制，识别疾病中的缺陷途径，并利用这些知识开发新的抗炎疗法。这项工作将大大有利于关节炎和肌肉骨骼治疗领域。

DESCRIPTION (provided by applicant): Skin inflammation involves local production of cytokines, which mediate recruitment of leucocytes and altered gene expression and cellular phenotypes of keratinocytes and other cells residing in the skin. lnterleukin-1 (IL-1) is an important mediator of local and systemic inflammation. IL-1 acts via a receptor complex involving the IL-1 receptor type I (IL-1RI) and membrane bound IL-1 receptor accessory protein (mlL-1 RAcP). A soluble isoform of IL-1 RAcP (slL-1 RAcP) appears to be an inhibitor of IL-1 signaling. The Principal Investigator has identified a novel soluble isoform of IL-1 RAcP, slL-1RAcP-(3, which has a unique C-terminus. Recently discovered cytokines, related to IL-1, appear to have activities similar to IL-1, but act via a different membrane receptor, IL-1 receptor related protein 2 (IL-1Rrp2). IL-1Rrp2 is primarily expressed in epithelial tissues, and preliminary data suggests that different IL-1 family members orchestrate distinct immune responses by inducing distinct cytokine expression profiles in primary human keratinocytes. Interestingly, IL-1 Rrp2 also utilizes mlL-1 RAcP for intracellular signaling. The primary hypothesis is that the two different isoforms of soluble IL-1 RAcP, slL-1 RAcP and slL-1 RAcP-|3, may inhibit the activities of all IL-1 family members or a sub-set of these. Furthermore, proportional expression of the three different isoforms of IL-1 RAcP may determine cellular responses to inflammatory stimuli. Since IL-1Rrp2 is primarily expressed in epithelial cells the involved mechanism may play a significant role in skin homeostasis and inflammation. ecombinant soluble IL-1 RAcP proteins will be expressed and purified. Human keratinocytes will be treated with agonist IL-1 family members and activation of gene expression will be examined using quantitative reverse transcription PCR and ELISA assays. Activities of the two soluble IL-1 RAcP isoforms will be examined in co-treatment experiments with individual cytokines. Expression of the three different IL-1 RacP isoforms and the IL-1 family members in keratinocytes will be determined following treatments with known modulators of IL-1 RAcP proportional expression and members of the IL-1 family. This project will provide novel information about the function(s) of the novel IL-1 related cytokines and the two soluble IL-1 RacP isoforms. The ultimate goals are to elucidate molecular mechanisms regulating inflammation, identify defective pathways in diseases and utilize this knowledge to develop novel anti-inflammatory therapies. of this work would greatly benefit arthritic and musculoskeletal therapeutic areas.