梅毒螺旋体对人脑微血管内皮细胞趋化因子配体6、8、10表达的影响

doi:10.3760/cma.j.issn.0412-4030.2018.05.008

摘要/Abstract

摘要： 目的探讨梅毒螺旋体对人脑微血管内皮细胞（HBMEC）趋化因子配体（CXCL）表达的影响。方法将培养的HBMEC分为4组，分别加入用细胞培养液稀释的有活性的梅毒螺旋体、灭活的梅毒螺旋体、脂多糖和细胞培养液（梅毒螺旋体组、灭活梅毒螺旋体组、脂多糖组和空白对照组）刺激6、12、24 h，用荧光定量PCR和酶联免疫吸附试验分别检测细胞中CXCL6、CXCL8和CXCL10基因和蛋白表达水平。利用Transwell小室细胞迁移实验检测梅毒螺旋体刺激后HBMEC对人早幼粒细胞白血病细胞HL-60的趋化作用。结果在各时间点，梅毒螺旋体组HBMEC中CXCL6、CXCL8和CXCL10的mRNA表达水平均显著高于空白对照组和灭活梅毒螺旋体组（P＜0.05），而灭活梅毒螺旋体组与空白对照组之间差异无统计学意义（均P＞0.05）。和脂多糖组相比，梅毒螺旋体组的CXCL6和CXCL8 mRNA表达降低（P＜0.05），CXCL10差异无统计学意义（P＞0.05）。在各时间点，梅毒螺旋体组HBMEC培养上清液中CXCL6和CXCL8含量均高于空白对照组和灭活梅毒螺旋体组（均P＜0.05），而后2组间差异无统计学意义（均P＞0.05）。各时间点培养上清液中CXCL10含量在梅毒螺旋体组、灭活梅毒螺旋体组和空白对照组间差异无统计学意义（均P＞0.05）；梅毒螺旋体组Transwell小室下室中迁移的HL-60细胞数量（A570）均高于另2组（均P＜0.05）。结论有活性的梅毒螺旋体可上调HBMEC中CXCL6、CXCL8和CXCL10基因表达水平，增加CXCL6和CXCL8的分泌，增强HBMEC对HL-60细胞的趋化作用，这可能在神经梅毒的发病中起一定作用。

关键词: 神经梅毒, 苍白密螺旋体, 趋化因子类, 细胞迁移分析, 白细胞, 人脑微血管内皮细胞, HL-60细胞

Abstract: Wu fan, Hu Wenlong, Xu Bufang, Wang Qianqiu Department of Sexually Transmitted Disease Clinical Management, Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College, Nanjing 210042, China （current affiliation of the first author was Department of Dermatology, Sir Run Run Hospital, Nanjing Medical University, Nanjing 211100, China） Corresponding author: Wang Qianqiu, Email: wangqq@ncstdlc.org 【Abstract】 Objective To evaluate the effects of Treponema pallidum （T. pallidum） on the of chemokine ligands （CXCL） in human brain microvascular endothelial cells（HBMECs）. Methods HBMECs were randomly divided into 4 groups, which were treated with viable T. pallidum suspension （T. pallidum group）, heat-inactivated T. pallidum suspension （inactivated T. pallidum group）, 200 μg/L lipopolysaccharide （LPS group） and cell culture medium （blank control group）, respectively, for 6, 12 and 24 hours. Fluorescence-based quantitative PCR and enzyme-linked immunosorbent assay （ELISA） were performed to determine the mRNA and protein of CXCL6, CXCL8 and CXCL10 in HBMECs in the above groups respectively. Transwell migration assay was conducted to evaluate the effects of T. pallidum-stimulated HBMECs on the chemotaxis of human promyelocytic HL-60 leukemia cells（HL-60 cells）. Results At 6, 12 and 24 hours, the T. pallidum group showed significantly higher mRNA of CXCL6, CXCL8 and CXCL10 in HBMECs compared with the blank control group and inactivated T. pallidum group （all P < 0.05）, while there were no significant differences between the blank control group and inactivated T. pallidum group （all P > 0.05）. Compared with the LPS group, the T. pallidum group showed significantly decreased mRNA of CXCL6 and CXCL8（P < 0.05）, but similar mRNA of CXCL10（P > 0.05）at 6, 12 and 24 hours. At these time points, the levels of CXCL6 and CXCL8 in the culture supernatant of HBMECs were significantly higher in the T. pallidum group than in the blank control group and the inactivated T. pallidum group （all P < 0.05）, but no significant differences were observed between the blank control group and the inactivated T. pallidum group （both P > 0.05）. Moreover, there were no significant differences in the level of CXCL10 in the culture supernatant of HBMECs among the T. pallidum group, the inactivated T. pallidum group and the blank control group （all P > 0.05）. The number of migratory HL-60 cells in the lower Transwell chambers was significantly higher in the T. pallidum group than in the inactivated T. pallidum group and the blank control group （both P < 0.05）. Conclusion Viable T. pallidum can up-regulate the gene of CXCL6, CXCL8 and CXCL10 in HBMECs, promote the secretion of CXCL6 and CXCL8, and enhance the chemotactic effect of HBMECs on HL-60 cells, which may play a certain role in the occurrence of neurosyphilis.

Key words: Neurosyphilis, Treponema pallidum, Chemotactic factors, Cell migration assays, leukocyte, Human brain microvascular endothelial cells, HL-60 cells

吴凡胡文龙许卜方王千秋. 梅毒螺旋体对人脑微血管内皮细胞趋化因子配体6、8、10表达的影响[J]. 中华皮肤科杂志, 2018, 51(5): 358-362.

Fan Wu. Effects of Treponema pallidum on the of chemokine ligand 6, 8, 10 in human brain microvascular endothelial cells[J]. Chinese Journal of Dermatology, 2018, 51(5): 358-362.

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