尿酸刺激树突状细胞成熟与Toll样受体mRNA表达关系的研究

目的 观察未成熟树突状细胞（DC）经尿酸刺激成熟过程中，TLRs mRNA的表达情况。方法 分离培养小鼠骨髓来源未成熟 DC。以尿酸体外刺激48 h后，RT-PCR方法检测DC TLR2 mRNA、TLR3 mRNA、TLR4 mRNA 、IL-12p70 mRNA等的表达，ELISA法检测上清IL-12P70水平。尿酸及NF-κB抑制剂PDTC刺激不同时间点时，免疫印迹法检测DC NF-κB p65活化情况。结果 尿酸刺激后DC TLRs mRNA 表达出现明显的变化，与培养基对照组相比，TLR2 mRNA、TLR3 mRNA、TLR4 mRNA表达下降（P＜0.05），以TLR4 mRNA下降最明显；IL-12p70表达显著增高（P＜0.05）；与LPS组相比，TLRs mRNA 与IL-12p70表达水平相似（P＞0.05）。尿酸刺激后15～45 min，DC核因子NF-κB p65显著活化。结论 尿酸能调节未成熟树突细胞 TLR2 mRNA、TLR3 mRNA、TLR4 mRNA及IL-12 p70的表达，促进NF-κB向核内转移，促进DC成熟。TLRs mRNA的动态变化可能为其诱导 DC 成熟及免疫功能提高的机制之一。

Expression of Toll like receptors mRNA study during the maturation of dendritic cells simulated by uric acid

Ma Xiao-jun1☆,Chen Xiao-ping1, Tian De-ying2 (1☆Department of Infectious Disease, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou 510300, Guang Dong, China, E-mail:mxjzjl@163.com,TEL:020-83827812-50110) OBJECTIVE: To investigate the expression of Toll like receptors mRNA in dendritic cells afer simulating with uric acid. METHODS: DCs were obtained from murine bone-marrow and cultured in vitro. After treatment with uric acid for 48 hours, The total RNA of DCs were extracted, mRNA expression of TLR2, TLR3, TLR4 and IL-12p70 in DCs was performed by RT-PCR method. After immature DCs were stimulated with uric acid (200μg) and NF-κB inhibitor PDTC for 15, 30 or 45 min, cytoplasmic or nuclear extracts were collected and were subjected to immunoblot analysis with Abs specific for NF-κB p65. RESULTS: The expression of TLR2 mRNA、TLR3 mRNA、TLR4 mRNA reduced significantly in DC treated with uric acid (200μg/ml)( P＜0.05) . Uric acid could increase the IL-12p70 mRNA expression of DC (P＜0.05). The effect of uric acid was similar with LPS stimulation on DC (P＜0.05). Increased expression of NF-κB p65 in DCs nuclear, within 15 min of uric acid conditioning of immature DC. In 30 min, the peak of these proteins was appear. Treatment of DC with PDTC, blocked the expression of NF-κBp65, in response to uric acid stimulation. CONCLUSION: Uric acid could down-regulate the mRNA expressions of TLR2, TLR3, TLR4, and enhance the expression of NF-κB, up-regulate the expressions of IL-12 p70 mRNA. Regulation of TLRs expressions may be one of the maturation mechanism of dendritic cells simulating by uric acid.