2021 年第 5 期 第 16 卷

慢性间歇低氧对小鼠巨噬细胞补体C3表达及细胞坏死的影响

Effect of chronic intermittent hypoxia on complement C3 expression and cell necrosis in mouse macrophages

作者：王越1吴铮1王悦1刘倍倍1陈蕾蕾1王志强1张慧娜2谢江3葛长江1蒋宏峰4吴小凡1

英文作者：Wang Yue1 Wu Zheng1 Wang Yue1 Liu Beibei1 Chen Leilei1 Wang Zhiqiang1 Zhang Huina2 Xie Jiang3 Ge Changjiang1 Jiang Hongfeng4 Wu Xiaofan1

单位：1首都医科大学附属北京安贞医院心内科北京市心肺血管疾病研究所100029；2首都医科大学附属北京安贞医院北京市心肺血管疾病研究所上气道功能障碍相关心血管疾病研究室100029；3首都医科大学附属北京安贞医院呼吸与危重症医学科100029；4首都医科大学附属北京安贞医院北京市心肺血管疾病研究所省部共建教育部心血管重塑相关疾病重点实验室100029

英文单位：1Department of Cardiology Beijing Anzhen Hospital Capital Medical University Beijing Institute of Heart Lung and Blood Vessel Diseases Beijing 100029 China; 2Upper Airway Dysfunction Related Cardiovascular Disease Research Laboratory Beijing Anzhen Hospital Capital Medical University Beijing Institute of Heart Lung and Blood Vessel Diseases Beijing 100029 China; 3Department of Respiratory and Critical Medical Beijing Anzhen Hospital Capital Medical University Beijing 100029 China; 4the Key Laboratory of Cardiovascular Remodeling-related Diseases Ministry of Education Beijing Anzhen Hospital Capital Medical University Beijing Institute of Heart Lung and Blood Vessel Diseases Beijing 100029 China

关键词：慢性间歇低氧；补体C3；巨噬细胞；坏死

英文关键词：Chronicintermittenthypoxia;ComplementC3;Macrophage;Necrosis

• 摘要：

• 目的 探讨慢性间歇低氧（CIH）对小鼠巨噬细胞补体C3表达及细胞坏死的影响。方法 采用小鼠单核巨噬细胞系RAW264.7细胞，以计算机控制低氧培养箱内氧浓度在5.0%和21.0%之间循环以建立CIH模型，并根据是否建立CIH模型分为低氧组和常氧组。在实验48、60、72 h时以实时荧光定量聚合酶链反应（RT-PCR）检测低氧诱导因子1α（HIF-1α）、补体C3、补体C3a受体（C3aR）和受体相互作用蛋白激酶3（RIP3）mRNA水平，蛋白质印迹法检测HIF-1α和RIP3蛋白表达水平，总抗氧化能力检测试剂盒检测2组细胞总抗氧化能力。结果 实验60、72 h时低氧组HIF-1α mRNA水平均明显高于常氧组［(2.21±0.68)比(1.00±0.19)，(8.52±1.80)比(1.00±0.64)］（均P＜0.05），蛋白水平呈现同样趋势。实验60、72 h时低氧组补体C3和C3aR的mRNA水平均明显高于常氧组［补体C3：(1.67±0.21)比(1.00±0.10)，(1.71±0.18)比(1.00±0.16)；C3aR：(1.87±0.07)比(1.00±0.06)，(2.15±0.36)比(1.00±0.22)］（均P＜0.05）。实验72 h时，RT-PCR检测结果 显示低氧组RIP3 mRNA水平明显高于常氧组［(2.83±1.07)比(1.00±0.19)，P=0.043］，蛋白水平呈现同样趋势。与常氧组相比，低氧组实验72 h时巨噬细胞总抗氧化能力显著下降［(37±23)μmol/mg比(235±48)μmol/mg，P=0.003］。结论 CIH下存在补体激活和巨噬细胞坏死增加，活化补体C3可能是CIH介导巨噬细胞坏死的通路之一。

• Objective To investigate the effect of chronic intermittent hypoxia (CIH) on complement C3 expression and cell necrosis in mouse macrophages. Methods The mouse mononuclear macrophage cell line RAW264.7 cells were used to generate CIH model by cycling oxygen concentration at 5.0%-21.0% within a computer-controlled hypoxia incubator. According to whether the CIH model was established, they were divided into hypoxia group and normoxia group. Real-time fluorescence quantitative polymerase chain reaction(RT-PCR) was used to detect the mRNA levels of hypoxia inducible factor-1α(HIF-1α), complement C3, complement C3a receptor (C3aR) and receptor interacting protein kinase 3(RIP3) at the 48, 60 and 72 h of the experiment. Western blotting was used to detect the protein expressions of HIF-1α and RIP3. Total antioxidant capacity assay kit was used to detect the total antioxidant capacity of cells in the two groups. Results The levels of HIF-1α mRNA in hypoxia group were significantly higher than those in normoxia group at 60 and 72 h of the experiment［(2.21±0.68) vs (1.00±0.19), (8.52±1.80) vs (1.00±0.64)］(both P＜0.05), and the protein expressions showed the same trend. Complement C3 and C3aR mRNA levels at 60 and 72 h of the experiment in hypoxia group were significantly higher than those in normoxia group［complement C3: (1.67±0.21) vs (1.00±0.10)，(1.71±0.18) vs (1.00±0.16); C3aR: (1.87±0.07) vs (1.00±0.06)，(2.15±0.36) vs (1.00±0.22)］（all P＜0.05）. At the 72 h of the experiment, RT-PCR showed that the level of RIP3 mRNA in hypoxia group was significantly higher than that in normoxia group［(2.83±1.07) vs (1.00±0.19), P=0.043］, and the protein level showed the same trend. Compared with normoxia group, the total antioxidant capacity of macrophages in hypoxia group significantly decreased at 72 h of the experiment［(37±23)μmol/mg vs (235±48)μmol/mg, P=0.003］. Conclusions Under CIH, there are complement activation and macrophage necrosis increase. Activation of complement C3 may be one of the pathways of CIH-mediated macrophage necrosis.