中华皮肤科杂志 ›› 2009, Vol. 42 ›› Issue (7): 481-483.

• 论著 • 上一篇    下一篇

CpG ODN刺激的寻常性银屑病中性粒细胞培养上清液对HaCaT细胞增殖的影响

宋珺1,潘萌2,罗邦国3,郑捷2,等4   

  1. 1.
    2. 上海交通大学医学院附属瑞金医院皮肤科
    3. 上海市第二医科大学附属瑞金医院皮肤科
    4. 湖北省中山医院整形美容外科
  • 收稿日期:2008-06-11 修回日期:2009-04-09 出版日期:2009-07-15 发布日期:2009-07-08
  • 通讯作者: 宋珺

Effect of culture supernatant of CpG ODN-stimulated neutrophils from patients with psoriasis vulgaris on the proliferation of keratinocytes

  • Received:2008-06-11 Revised:2009-04-09 Online:2009-07-15 Published:2009-07-08

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摘要：

目的 探讨中性粒细胞在银屑病发病中的作用。方法 采用脂多糖（LPS）或人工合成的CpG ODNs（CpG-A或CpG-B）刺激中性粒细胞，取其培养上清液处理人永生化角质形成细胞株（HaCaT），观察HaCaT细胞株增殖的变化，并通过抗IL-8、抗TNF-α单克隆抗体以及SOD/CAT预处理评价中性粒细胞分泌的炎性因子在银屑病发病中的作用。结果 与RPMI 1640培养液比较，无LPS、CpG-A和CpG-B刺激的正常人和静止期银屑病患者中性粒细胞培养上清液对HaCaT细胞株的增殖无明显促进作用（P > 0.05），而进行期患者中性粒细胞培养上清液明显促进HaCaT细胞株的增殖（t = 2.41，P < 0.05）。与正常人比较，进行期银屑病患者中性粒细胞受LPS、CpG-A和CpG-B刺激后的培养上清液显著增强HaCaT细胞的增殖（t值分别为3.11，2.89，2.29，P < 0.05）。经LPS、CpG-A刺激的中性粒细胞培养上清液与无刺激组比较，均明显促进HaCaT细胞的增殖。与未予阻断剂组比较，予抗IL-8、抗TNF-α单克隆抗体以及SOD/CAT预处理的进行期银屑病组中性粒细胞经LPS、CpG-A和CpG-B刺激后的上清液诱导的HaCaT细胞增殖均显著减少（P < 0.05）。结论 寻常性银屑病患者外周血中性粒细胞存在炎性因子IL-8、TNF-α和SOD/CAT分泌异常，且这些异常分泌的炎性因子可以促进HaCaT细胞的增殖。

关键词: 寻常型银屑病，中性粒细胞，角质形成细胞

Abstract:

Objective To investigate the role of neutrophils in the pathogenesis of psoriasis vulgaris. Methods Neutrophils were isolated from venous blood samples of 25 patients with psoriasis vulgaris （including 13 cases of active psoriasis and 12 cases of inactive psoriasis） as well as 25 normal human controls, and cultured. Then, these neutrophils were grouped and treated with lipopolysaccharide （LPS, 100 g/L）, CpG-A （50 mg/L）, CpG-B （50 mg/L）, and RPMI 1640 culture medium, respectively, for 24 hours followed by the collection of culture supernatants. Human keratinocytes （HaCaT） were cultured in the presence of supernatants of treated or untreated neutrophils for 72 hours followed by the detection of cell proliferation with MTT assay. To determine the role of proinflammatory factors, SOD/CAT and monoclonal antibody to IL-8 and TNF-alpha of 400 u/mL were used to pretreat HaCaT cells 1 hour prior to the stimulation with supernatants of neutrophils. Results Compared with culture medium, the supernatant of unstimulated neutrophils from normal controls or patients with inactive psoriasis had no significant effect on the proliferation of HaCaT cells （P > 0.05）, but that from patients with active psoriasis markedly promoted the proliferation of HaCaT cells （t = 2.41, P < 0.05）. After stimulation by LPS, CpG-A and CpG-B, the supernatant of active patient- derived neutrophils significantly promoted the proliferation of HaCaT cells compared with that of normal control-derived neutrophils （t = 3.11, 2.89, 2.29, respectively, all P < 0.05）. In comparison with unstimulated neutrophils, the supernatant from LPS- and CpG-A stimulated neutrophiles significantly accelerated the proliferation of HaCaT cells. Furthermore, the proliferation of HaCaT cells induced by the supernatants of LPS-, CpG-A-, CpG-B-stimulated neutrophils from psoriatic patients was statistically suppressed by the pretreatment with the monoclonal antibody to IL-8, TNF-alpha and SOD/CAT （all P < 0.05）. Conclusions In patients with psoriasis vulgaris, there is an abnormal secretion of IL-8, TNF-alpha and superoxide by neutrophils in peripheral blood, and these proinflammatory factors could promote the proliferation of HaCaT cells.