The NLR family pyrin domain-containing 3 (NLRP3) inflammasome is a cytoplasmic multimolecular complex that generates interleukin (IL)-1β and is considered a main pathogenic mechanism for uric acid-induced inflammation. Whether toll-like receptor 9 (TLR9) is responsible for uric acid-induced NLRP3 inflammasome activation remains unclear. Thus, the aim of this study was to identify the role of TLR9 in NLRP3 inflammasome activation through monosodium urate (MSU) crystal-induced mitochondrial DNA. RAW 264.7 cells treated with MSU crystals, CpG oligonucleotides (ODNs), or a combination of both were used to assess nuclear factor (NF)-κB signaling, NLRP3 inflammasome components such as NLRP3, ASC, and caspase-1, and IL-1β. Real-time polymerase chain reaction (RT-PCR), Western blotting, DNA fragmentation assay, mitochondrial DNA copy number assay, and immunofluorescence were used in the in vitro study. RAW 264.7 cells treated with CpG-ODN stimulated the activation of NF-κB signaling, the NLRP3 inflammasome components NLRP3, ASC, and caspase-1, and IL-1β gene and protein expression. DNA fragmentation assay showed that MSU crystals induced cellular apoptosis. Fragmented DNA prompted by MSU crystals induced TLR9 expression. RAW 264.7 cells treated with CpG-ODN or MSU crystals and both increased expression of mitochondrial DNA relative to nuclear DNA. CpG-ODN and MSU crystals augmented the activation of NLRP3 inflammasome components and IL-1β expression, which was significantly suppressed in RAW 264.7 cells transfected with TLR9 siRNA. This study suggests that TLR9 activated by MSU crystal-mediated mitochondrial DNA contributes to the activation of NLRP3 inflammasomes and IL-1β production.

NLR 家族 pyrin 结构域 3 (NLRP3) 炎性体是一种细胞质多分子复合物，可产生白细胞介素 (IL)-1β，被认为是尿酸诱导炎症的主要致病机制。Toll 样受体 9 (TLR9​​) 是否负责尿酸诱导的 NLRP3 炎性体激活仍不清楚。因此，本研究的目的是通过尿酸单钠 (MSU) 晶体诱导的线粒体 DNA 确定 TLR9 在 NLRP3 炎症小体激活中的作用。用 MSU 晶体、CpG 寡核苷酸 (ODN) 或两者的组合处理的 RAW 264.7 细胞用于评估核因子 (NF)-κB 信号传导、NLRP3 炎性体成分，例如 NLRP3、ASC 和 caspase-1，以及 IL-1β . 实时聚合酶链反应 (RT-PCR)、蛋白质印迹、DNA 片段化测定、线粒体 DNA 拷贝数测定、体外研究。用 CpG-ODN 处理的 RAW 264.7 细胞刺激 NF-κB 信号传导、NLRP3 炎性体成分 NLRP3、ASC 和 caspase-1 以及 IL-1β 基因和蛋白质表达的激活。DNA片段化测定表明MSU晶体诱导细胞凋亡。由 MSU 晶体引起的片段化 DNA 诱导 TLR9 表达。用 CpG-ODN 或 MSU 晶体处理的 RAW 264.7 细胞都增加了线粒体 DNA 相对于核 DNA 的表达。CpG-ODN 和 MSU 晶体增强了 NLRP3 炎性体成分的激活和 IL-1β 表达，这在用 TLR9 siRNA 转染的 RAW 264.7 细胞中被显着抑制。该研究表明，由 MSU 晶体介导的线粒体 DNA 激活的 TLR9 有助于 NLRP3 炎性体的激活和 IL-1β 的产生。