OCILRP2-Fc抑制小鼠骨髓来源的树突状细胞分化成熟

OCILRP2-Fc inhibits maturation of murine bone marrow-derived dendritic cells

目的:采用OCILRP2胞外段蛋白OCILRP2-Fc阻断OCILRP2受体,探讨OCILRP2对小鼠树突状细胞发育成熟的影响及其机制。方法:分离培养小鼠骨髓来源的树突状细胞(BMDC),荧光定量RT-PCR和流式细胞术分析OCILRP2在不成熟BMDC和LPS诱导的成熟BMDC的表达差异;流式细胞术分析OCILRP2-Fc对BMDC细胞表面标志分子的表达以及对BMDC抗原摄取能力的影响;ELISA检测培养上清中细胞因子的浓度,分析OCILRP2-Fc对BMDC分泌炎性细胞因子的影响;Western blot检测转录因子NF-κB的活化,分析OCILRP2-Fc影响BMDC分化成熟的分子机制。结果:荧光定量RT-PCR和流式细胞术结果均表明LPS可以显著上调OCILRP2在BMDC的表达(P<0.05);与对照组相比,OCILRP2-Fc可以明显抑制MHCⅡ类分子及共刺激分子CD80在LPS诱导的成熟BMDC的表达;和成熟BMDC相比,OCILRP2-Fc组BMDC经LPS诱导24 h后仍然具有较强的抗原吞噬能力;而且,OCILRP2-Fc组BMDC培养上清中炎性细胞因子IL-6、IL-12、TNF-α的浓度明显低于对照组(P<0.05)。Western blot结果表明OCILRP2-Fc抑制了LPS诱导的I-κB降解和NF-κB p65的磷酸化。结论:OCILRP2-Fc阻断OCILRP2受体信号,通过抑制LPS诱导的NF-κB活化影响小鼠BMDC分化成熟。提示OCILRP2受体在树突状细胞的分化成熟过程中具有促进作用。

Objective：Osteoclast inhibitory lectin-related protein 2（OCILRP2） blocked by recombinant extracellular domain protein OCILRP2-Fc,to investigate the role of OCILRP2 plays on murine bone marrow derived-dendritic cells（ BMDC）maturation. Methods： BMDC were obtained from BALB / c mouse bone marrow. Real-time PCR and flow cytometry were performed for detecting the different OCLRP2 expression between immature BMDC and LPS induced mature BMDC. The cell surface markers expression and antigen uptake were analyzed by flow cytometry. ELISA was carried on measuring the proinflammatory cytokine concentration in cell culture supernatant. NF-κB activation were analyzed by Western blot to explore the molecular mechanism of OCILRP2-Fc effect on BMDC maturation. Results： Relative quantitative RT-PCR and flow data showed OCILRP2 expression in LPS induced mature BMDC was significantly elevated（ P 〈0. 05）. In contrast to control group,OCILRP2-Fc obviously downregulated MHC-Ⅱ and co-stimulator CD80 expression on LPS induced mature BMDC. Moreover,LPS induced mature BMDC with OCILRP2-Fc treatment still have strong antigen uptaking capability. And the cytokine concentration of IL-6,IL-12,TNF-α in BMDC culture supernatant were significantly decreased by OCILRP2-Fc treatment（ P 〈0. 05）. Western blot data showed that OCILRP2-Fc restrained LPS induced I-κB degradation and NF-κB p65 phosphorylation in BMDC. Conclusion： OCILRP2-Fc inhibits LPS induced BMDC maturation by blocking OCILRP2 signaling and restraining NF-κB activation,which suggests OCILRP2 plays important roles in BMDC maturation.