特异性miRNAs在急性痛风性关节炎中的作用机制及白藜芦醇干预效果研究

NLRP3 inflammasome and chemokines play vital roles in the pathogenesis of acute gout arthritis (AG). While microRNAs (miRNAs) are important regulatory factors in many inflammatory diseases, their role in AG remains poorly understood. Our preliminary studies have indentified three gout-specific miRNAs including miR-223-3p, miR-22-3p and miR-374b-5p that may target NLRP3 inflammasome or chemokine ligand 2 (CCL2) (predicted by bioinformatics analyses) and express at significantly lower levels in AG compared to chronic hyperuricaemia. On the other hand, resveratrol inhibits the expression of NLRP3 inflammasome and chemokines in AG. Hereby we hypothesize that these miRNAs may inhibit AG pathogenesis via regulating NLRP3 and chemokines, while resveratrol may inhibit the expression of inflammatory proteins via regulating miRNAs. In the proposed study, using cell culture and an AG rat model, we will investigate the the regulatory roles of these three gout-specific microRNAs in acute gouty arthritis and the intervention effect of resveratrol at cellular and systemic levels by a variety of techniques including real-time PCR, adenovirus transfection and intra-articular injection of miRNA mimics. This study will extend our understanding on the pathogenesis of AG in terms of epigenetics, and provide insight for development of new targeted treatments for gout.

NLRP3炎性体和趋化因子在急性痛风性关节炎(AG)发病中有重要作用。miRNA在多种炎症性疾病中有关键调控作用，但其与AG的关系仍知之甚少。我们前期研究发现了可能靶向NLRP3或趋化因子CCL2并在AG中呈特异性低表达的miR-223-3p、miR-22-3p和miR-374b-5p；白藜芦醇在AG中显著降低NLRP3和趋化因子的表达。为此，我们提出假说：这些miRNAs在AG中可靶向调控NLRP3或CCL2的表达从而抑制AG发病；而白藜芦醇可通过调控miRNA表达水平方式抑制炎症相关蛋白表达。为了验证这一假说，我们将通过体外细胞培养及AG动物模型，采用实时定量PCR、腺病毒转染、关节腔内注射miRNA等手段，从细胞、组织及动物整体水平等多方面探讨上述痛风特异性miRNA在AG中的作用机制及白藜芦醇的干预效果。本研究将在表观遗传学层面阐述AG发病机制，并为研发新的治疗手段提供理论基础。

NLRP3炎性体在痛风性关节炎发病中有重要作用。MicroRNA在多种炎症性疾病中有关键调控作用，其与痛风性关节炎的关系仍需阐明。本研究旨在探究miRNAs对痛风患者，单核巨噬细胞以及动物模型中炎性因子的调节作用。于是我们结合相关临床患者进行了统计分析，同时构建了THP-1细胞炎症模型，并给与尿酸盐结晶（MSU）刺激，通过蛋白质印记法、实时荧光定量核酸扩增检测、酶联免疫吸附测定实验技术分析miRNAs以及各性因子在炎症中的表达变化，通过过表达或抑制miRNAs来观察其与各炎性因子的相关性。随后通过双荧光报告实验验证miRNA223-3p及miRNA22-3p的炎症调控靶标，确定miRNAs与炎症因子的调控关系。同时探究白藜芦醇与miRNAs共同的的抗炎作用。最后通过蛋白质印记法、实时荧光定量核酸扩增检测、酶联免疫吸附测定、免疫组织化学染色对小鼠痛风气囊模型进行验证。我们发现miRNA223-3p与miRNA22-3p可通过靶向调控NLRP3炎性小体的表达，起到减轻MSU诱导的痛风性关节炎炎症反应的作用，并且白藜芦醇与miRNA联合作用可以更加有效的抑制炎症反应。本研究将对急性痛风性关节炎的治疗提供新的治疗方向。