CsNOSIP与NLRP3炎症小体活化的关系及其在华支睾吸虫慢性感染过程中的作用

The relationship of CsNOSIP and NLRP3 inflammasome activation and the potential role of CsNOSIP in the chronic infection process of Clonorchis sinensis

目的探讨华支睾吸虫一氧化氮合酶相互作用蛋白(CsNOSIP)与Nod样受体蛋白3(NLRP3)炎症小体活化的关系及其在华支睾吸虫慢性感染过程中的作用。方法脂多糖(LPS)和三磷酸腺苷(ATP)共同刺激构建NLRP3炎症小体活化人巨噬细胞(THP-1)细胞模型,荧光定量PCR(qRT-PCR)检测不同浓度CsNOSIP作用下,NLRP3炎症小体相关指标NLRP3和白细胞介素-1b(IL-1b)的表达情况;qRT-PCR法检测CsNOSIP直接刺激人肝星状细胞LX-2后肝纤维化相关指标的表达情况;CsNOSIP与华支睾吸虫溶血磷脂酶(CsLysoPLA)共孵育后,qRT-PCR法检测小鼠巨噬细胞RAW264.7白细胞介素-25(IL-25)的分泌情况;构建RAW264.7炎症细胞模型,qRT-PCR法检测肿瘤坏死因子-α(TNF-α)的表达情况。结果 qRT-PCR法检测结果显示,低浓度的CsNOSIP可抑制THP-1细胞模型内NLPR3炎症小体相关指标NLRP3和IL-1b的表达,与阳性对照组相比差异有统计学意义(P<0.05)。并且CsNOSIP还能下调CsLysoPLA诱导的IL-25的表达,与单独CsLysoPLA相比,差异有统计学意义(P<0.05)。CsNOSIP还可抑制LPS诱导的TNF-α的表达(F=18.71,P<0.05)。结论 CsNOSIP可抑制NLRP3炎症小体活化及下调IL-25和TNF-α的表达,可能在维持华支睾吸虫慢性感染的过程中发挥着重要的作用。

Objective To explore the effect of Nitride oxide synthase interacting proteins（CsNOSIP）on the activation of Nod-like receptor protein 3（NLRP3）inflammasome and its potential role in keeping chronicity of Clonorchis sinensis.Methods NLRP3 inflammasome activation of THP-1 cells model was established by lipopolysaccharide（LPS）and adenosine triphosphate（ATP）,to detect the mRNA levels of inflammasome related-genes-NLRP3 and interleukin-1 b（IL-1 b）after treated with different concentrations of CsNOSIP. The LX-2 cell line was treated with different concentrations of CsNOSIP and liver fibrosis related markers were detected by qRT-PCR. RAW264.7 cell line was co-stimulated by CsNOSIP and Lysophospholipase A（CsLysoPLA）to study the effect of CsNOSIP on the expression of interleukin-25（IL-25）induced by CsLyso PLA. LPS-induced inflammatory RAW264.7 cell model was established to detect the mRNA level of tumor necrosis factor-α（TNF-α）after co-cultured with CsNOSIP. Results According to the results of qRT-PCR, the expression of NLRP3 inflammsome related genes-NLRP3 and IL-1 b in THP-1 cells were down regulated by low dose of CsNOSIP when compared with the positive group（P〈0.05）. Moreover,CsNOSIP could down regulate the m RNA levels of IL-25 induced by CsLysoPLA （P〈0.05）,and TNF-α induced by LPS（F=18.71,P〈0.05）. Conclusion CsNOSIP could suppress the activation of NLRP3 inflammasome and probably play important role in keeping long-standing infection of Clonorchis sinensis.