Innate Immunity in gouty Inflammation

描述(申请人提供)：在痛风患者中，尿酸单钠(MSU)晶体沉积在关节中会刺激急性中性粒细胞炎症和慢性滑膜炎，可能导致骨和软骨破坏。MSU晶体引发炎症在很大程度上是通过它们直接激活细胞的能力，包括滑膜衬里细胞和吞噬细胞。然而，惰性MSU晶体诱导痛风炎症的细胞识别因子的确切分子同一性尚不清楚。我们最近发现，天然免疫模式识别受体(PRRs)的质膜TLR2、TLR4及其共同的细胞内适配蛋白MyD88在体外介导了MSU晶体惰性、无内毒素制剂的细胞识别，并在体内决定了MSU晶体的炎症潜能。此外，我们观察到另一种PRR CD14，即TLR2和TLR4辅助分子，在体外直接与MSU晶体结合，并在体内介导MSU晶体诱导的炎症反应。此外，我们现在观察到，细胞内的PrR低温比林/NALPS在促进MSU晶体诱导的体内炎症中也发挥着重要作用。因此，本研究的中心目标是检验这样一种假设，即特定的细胞外(CD14、TLR2和TLR4)和细胞内(低温比林/NALP3)的天然免疫PRR的桥接协调地转导和联系细胞外的参与和摄取与细胞内对MSU晶体的反应，而MSU晶体是痛风炎症的中心。要做到这一点，我们将在两个目标上检验这一假设。在目标1中，我们将验证CD14、TLR2和TLR4与MSU晶体的胞外结合是MSU晶体诱导吞噬细胞炎症反应所必需的假设。在目标2中，我们将验证两种不同的细胞内天然免疫机制控制MSU晶体诱导的炎症反应的假设：(1)规范的TLR2和TLR4信号通过TLR接头蛋白MyD88导致NF-kappaB调节的炎症细胞因子的表达。(2)依赖内化的(MSU晶体结合的)CD14、TLR2或TLR4的低温比林/NALPS炎性小体途径依赖于MyD88非依赖的细胞内参与介导IL-1β的成熟和血清。这项工作的完成将有助于进一步了解急性痛风炎的机制，如何控制痛风性关节炎和潜在的其他形式的晶体诱导的炎症。

DESCRIPTION (provided by applicant): In gout, monosodium urate (MSU) crystals deposited in the joint stimulate both acute neutrophilic inflammation and chronic synovitis that may lead to bone and cartilage destruction. MSU crystals trigger inflammation in large part via their capacity to directly activate cells, including synovial lining cells and phagocytes. However, the precise molecular identity of the cell recognition factors for inert MSU crystals to induce gouty inflammation has remained unclear. We recently discovered that the innate immune pattern recognition receptors (PRRs) plasma membrane TLR2, TLR4, and their common intracellular adapter protein MyD88 mediate cellular recognition of inert, endotoxin-free preparations of MSU crystals in vitro, and determine the inflammatory potential of MSU crystals in vivo. In addition, we observed that another PRR CD14, the TLR2 and TLR4 accessory molecule, directly binds MSU crystals in vitro, and mediates MSU crystal-induced inflammatory responses in vivo. Moreover, we now observe that cryopyrin/NALPS, an intracellular PRR, also plays an important role in promoting MSU crystal-induced inflammation in vivo. Thus, the central objective of this study is to test the hypothesis that the bridging of specific extracellular (CD14, TLR2, and TLR4) and intracellular (cryopyrin/NALP3) innate immune PRRs coordinately transduce and link extracellular engagement and uptake to intracellular responses to MSU crystals, which are central to gouty inflammation. To do so, we will test the hypothesis in two aims. In aim 1, we will test the hypothesis that extracellular engagement of MSU crystals by CD14, TLR2, and TLR4 is essential for MSU crystal-induced inflammatory responses in phagocytes. In Aim 2, we will test the hypothesis that two distinct intracellular innate immune mechanisms govern inflammatory responses induced by MSU crystals: (1) canonical TLR2 and TLR4 signaling through the TLR adapter protein MyD88 leads to expression of NF-kappaB regulated inflammatory cytokines. (2) MyD88-independent intracellular engagement of the cryopyrin/NALPS inflammasome pathway dependent on internalized (MSU crystal-bound) CD14, TLR2 or TLR4 mediates IL-1beta maturation and seretion. Completion of this work will help further understanding the mechanisms of acute gouty inflammation, how to control gouty arthritis and potentially other forms of crystal-induced inflammation.