IL-10对Aa-LPS刺激的兔肺泡巨噬细胞功能的影响

Effects of IL-10 on Functions of Aa-LPS Induced Rabbit Alveolar Macrophages

目的研究白细胞介素10(IL-10)在伴放线放线杆菌(Aa)脂多糖(LPS)刺激兔肺泡巨噬细胞(AM)产生炎症反应过程中的免疫调节作用及其机制。方法分离纯化AM,100ng/mL IL-10预孵2h,加入Aa-LPS刺激继续培养24h,流式细胞术检测AM对荧光标记Aa(FITC-Aa)的吞噬指数(PI)情况及AM的活性氧(ROS)水平;荧光定量PCR法检测CD14、TLR4、SRA和MHC-Ⅱ基因表达的变化;硝酸还原酶法检测AM的NO分泌情况。结果 IL-10上调Aa-LPS诱导的AM的PI(P<0.05),下调Aa-LPS诱导的AM膜表面受体CD14、TLR4、MHC-ⅡmRNA的表达(P<0.01),而SRA mRNA表达量升高(P<0.05);IL-10抑制Aa-LPS诱导的AM的ROS和NO水平(P<0.01)。结论 IL-10可以通过上调Aa-LPS诱导的AM SRA基因表达,下调CD14、TLR4、MHC-Ⅱ基因表达,并且抑制其产生ROS和NO的水平,从而调节免疫反应,限制炎症介质的大量释放,同时增强AM的吞噬作用,清除感染。

Objective To evaluate the potential immunomodulatory effects and explore the possi ble mechanisms of interlukin 10 on activation of Actinobacillus actinomycetemcomitans （Aa）-LPS induced alveolar macrophages（AM）. Methods Isolation and purification of AM, AM was pre-incubated with 100 ng/mL IL-10 for 2 h and then activated by Aa LPS for 24 h： phagocytic index （PI） was measured by flow cytometry; RT-PCR was used to detect the expression of gene CD14, TLR4, SRA, MHC Ⅱ ; flow cytometry was used to assess ROS; nitrate reduetase assay was used to examin the amount of NO production. Results II.-10 increase the PI of Aa-LPS activated AM（P〈0.05）, remarkably inhibited the expressions of CD14, TLR4, MHC- Ⅱ （P〈0.01）, but enhanced the expression of SRA（P〈0.05）, significantly sup pressed the ROS（P〈0.01） and NO production（P〈0.01）. Conclusions IL-10 can modulate of immune response and down-regulate excessive inflammatory mediator production and enhance phagocytosis of Aa- LPS activated AM via increasing SRA, suppressing CD14, TLR4, MHC-Ⅱ expression, and inhibit ROS, NO production.