基础医学与临床 ›› 2016, Vol. 36 ›› Issue (3): 331-336.

• 研究论文 • 上一篇    下一篇

NO抑制体外IFN-γ刺激成肌细胞/肌管NLRP3炎性小体活化

刘幸卉1,张任飞2,史丹丹1,曹标1,术蓉1,谷瑞彩1,肖将尉1,廖华1   

  1. 1. 南方医科大学
    2. 四川省绵阳市第三人民医院
  • 收稿日期:2015-06-15 修回日期:2015-12-02 出版日期:2016-03-05 发布日期:2016-02-22
  • 通讯作者: 廖华 E-mail:hua-liao@163.com
  • 基金资助:
    国家自然科学基金面上项目;国家自然科学基金面上项目;广东省科技计划项目;广东省自然科学基金面上项目

Inhibition of nitric oxide on the activation of NLRP3 inflammasome myoblast /myotubes stimulated by IFN-γ in vitro

  • Received:2015-06-15 Revised:2015-12-02 Online:2016-03-05 Published:2016-02-22
  • Supported by:
    ; the Science and Technology Planning Project of Guangdong Province, People’s Republic of China

摘要: 目的 观察一氧化氮供体硝普钠(SNP)、一氧化氮合酶抑制剂(L-NAME)对体外炎性培养(IFN-γ刺激)成肌细胞/肌管NLRP3炎症小体活化的影响 方法 利用IFN-γ刺激小鼠C2C12成肌细胞/肌管,qPCR和Western blot分析炎性小体(ASC、NLRP3、caspase-1)的形成及活化、ELISA检测细胞培养上清IL-1β的分泌。进一步利用L-NAME和SNP处理IFN-γ诱导的C2C12细胞/肌管,分析炎性小体的形成、活化及IL-1β的分泌。结果 IFN-γ刺激培养后,成肌细胞/肌管中NLRP3、ASC和mature-caspase-1表达水平上调(P<0.01)。较之未刺激的细胞,IFN-γ诱导会显著上调成肌细胞/肌管培养基中的IL-1β浓度(P<0.01)。SNP处理后6h,分化肌管(IFN-γ刺激48h)内炎性小体(ASC、NLRP3、caspase-1)mRNA和蛋白水平较单纯刺激细胞显著下调(P<0.01),L-NAME则上调ASC、NLRP3、caspase-1表达水平(P<0.05)。与上述结果一致,SNP和L-NAME处理同时分别下调或上调培养基中IL-1β浓度(P<0.05)。结论 在炎性条件下,成肌细胞/肌管具备合成并活化NLRP3炎性小体的能力。NO对炎性小体的形成及活化有一定的抑制作用。

关键词: NO, C2C12细胞, 炎症小体

Abstract: Objective: To observe the effect of sodium nitroprusside (SNP) or nitro L arginine acid methyl ester (L-NAME) on the NLRP3 inflammasome activation of C2C12 myoblast or differentiated myotubes in inflammatory culture in vitro. Methods: C2C12 myoblasts were stimulated by IFN-γ, then the formation and activation of NLRP3 inflammasome were analyzed by qPCR and Western blot,and the secretion of IL-1β in the cell culture supernatant was detected by ELISA. Further, C2C12 myoblasts induced by IFN-γ were treated by L-NAME or SNP respectively, then the formation and activation of NLRP3 inflammasome and the secretion of IL-1β were analyzed. Results: qPCR and Western blotting detection confirmed that expression of NLRP3, ASC and mature-caspase-1 in C2C12 myoblasts or differentiation myotubes induced by IFN-γ was up-regulated(P<0.01). ELISA analysis further confirmed that the IL-1β concentration in the medium of C2C12 induced by IFN-γ was up-regulated compared with that of the non stimulating cells(P<0.01).At 6 h after SNP treatment, a significant down-regulation of mRNA and protein levels of differentiation myotube inflammasome (ASC and NLRP3, caspase-1) was found(P<0.01) and a contrary result was obtained after L-NAME treatment(P<0.05). Consisvent with the above results, the expression of IL-1βin the culture supernantent was down-regulated or up-regulated by SNP or L-NAME respectively(P<0.05). Conclusions: In inflammatory conditions, C2C12 myoblasts or differentiation myotubes possess the ability to synthesize inflammasome NLRP3. NO might cause a certain inhibition on the NLRP3 inflammasome formation and activation.

Key words: Nitric Oxide, C2C12 myoblast , Inflammasomes

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