主 办:北 京 中 医 药 大 学
ISSN 1006-2157 CN 11-3574/R

北京中医药大学学报 ›› 2017, Vol. 40 ›› Issue (4): 314-321.doi: 10.3969/j.issn.1006-2157.2017.04.010

• 科技之窗 • 上一篇    下一篇

益肺活血方对香烟提取物诱导巨噬细胞炎症反应的影响*

简芙平1, 欧敏1,2#   

  1. 1 南方医科大学第三临床医学院 广东 510515;
    2 海军总医院干部呼吸科
  • 收稿日期:2016-11-04 出版日期:2017-04-30 发布日期:2017-04-30
  • 通讯作者: 欧敏,女,博士,教授,硕士生导师,E-mail: oumin1999@aliyun.com
  • 作者简介:简芙平,男,在读硕士生
  • 基金资助:
    *军队中医药科研专项基金项目(No.10ZYZ219)

Effects of Yifei Huoxue Fang on the inflammatory response in macrophages induced by cigarette smoke extract*

JIAN Fuping1 OU Min1,2#   

  1. 1 The Third Clinical Medical School of Southern Medical University, Guangdong 510515;
    2 VIP Department of Respiration Disease, Navy General Hospital of PLA
  • Received:2016-11-04 Online:2017-04-30 Published:2017-04-30

摘要: 目的 探讨益肺活血方对香烟提取物诱导巨噬细胞炎症反应的影响。方法 用12-豆蔻酸-13-乙酸佛波醇将U937细胞诱导为巨噬细胞, 用1.2、2.4、4.8、9.6 g/L的益肺活血方及5、10、20、50、100 mL/L的香烟提取物处理巨噬细胞,于12、24、48 h用细胞增殖和毒性检测(CCK-8)法测定细胞活性。巨噬细胞分5组:正常细胞组、香烟提取物组、香烟提取物+益肺活血方低、中、高浓度组,酶联免疫吸附测定(ELISA)检测培养上清白细胞介素-8(IL-8)和肿瘤坏死因子-α(TNF-α)含量,RT-PCR法检测胞内IL-8 和TNF-α mRNA水平。机制实验分4组:正常细胞组、香烟提取物组、香烟提取物+益肺活血方(2.4 g/L)组及香烟提取物+NF-κB抑制剂组,蛋白印迹法(Western blot)检测胞内核转录因子-κB(NF-κB) p50、p65及p-IκB表达;免疫荧光法观察胞核内NF-κB p65及胞浆内p-IκB表达。结果 益肺活血方(9.6 g/L)作用细胞24、48 h后 及香烟提取物(100 mL/L)于各时间点均有细胞毒性(P<0.05)。与正常细胞组相比,香烟提取物组 IL-8及TNF-α表达上调,胞内NF-κB p50、p65及p-IκB表达亦升高(P<0.05),NF-κB核转位增加(P<0.05);益肺活血方可抑制诱导后的IL-8和TNF-α的表达以及NF-κB活性(P<0.05)。结论 益肺活血方可减少香烟提取物诱导的巨噬细胞表达IL-8 、TNF-α,抑制NF-κB活化可能是其机制之一。

关键词: 慢性阻塞性肺疾病, 益肺活血方, 巨噬细胞, 白细胞介素-8, 肿瘤坏死因子-α, 核转录因子-κB

Abstract: Objective To explore the effects of Yifei Huoxue Fang (YFHXF, lung-invigorating blood-activating formula) on the inflammatory response in macrophages induced by cigarette smoke extract. Methods U937 monocytic cells were differentiated into macrophages using phorbol 12-myristate 13-acetate (PMA). These PMA-induced macrophages were then treated with YFHXF at various concentrations (1.2,2.4,4.8,9.6 g/L) or exposed to cigarette smoke extract (CSE)(5,10,20,50,100 mL/L). At 12, 24, and 48 h, viability of the cells was assessed by using the cell counting kit-8 (CCK-8). Macrophages were divided into 5 groups: normal group, CSE group, CSE+ YFHXF low-dose (1.2 g/L) group, CSE+ YFHXF medium-dose (2.4 g/L) group, and CSE+ YFHXF high-dose (4.8 g/L) group. Levels of IL-8 and TNF-α in cell culture supernatant were detected by using enzyme-linked immunosorbent assay (ELISA), and gene expression of IL-8 and TNF-α were also measured with real-time polymerase chain reaction (RT-PCR). For the study of mechanism, macrophages were divided into 4 groups: normal group, CSE group, CSE+ YFHXF (2.4 g/L) group, and CSE+NF-κB inhibifant(parthenolide) group. The expression of nuclear factor-κB (NF-κB) p50/p65 and p-IκB was examined by using Western blot method. Meanwhile, the expression of nucleus p65 and plasma p-IκB in macrophages was also measured by using the immunofluorescent assay. Results Cytotoxicity was present in macrophages at all timing points induced by CSE (100 ml/L) and at 24, and 48 hours after treated with YFHXF (9.6 g/L). Compared with normal cell group, IL-8 and TNF-α of CSE group, and NF-κB p50/p65 and p-IκB expression were all upregulated (P<0.05). NF-κB nuclear translocation was increased (P<0.05). Yifei Huoxue Fang could inhibit the expression of IL-8 and TNF-α induced by CSE, and viability of NF-κB (P<0.05). Conclusion Yifei Huoxue Fang could inhibit the expression of IL-8 and TNF-α induced by CSE, possibly by inhibiting NF-κB activation.

Key words: chronic obstructive pulmonary disease, Yifei Huoxue Formula, macrophages, IL-8, TNF-α, nuclear factor-κB

中图分类号: 

  • R285.5