摘要
目的 :观察TLR1(Toll-like receptors 1)、TLR2和TLR6在卵巢癌患者外周血单个核细胞(peripheral blood mononuclear cells,PBMC)亚群中的表达水平,初步探索TLR1、TLR2和TLR6在卵巢癌发生发展中的作用机制。方法 :收集13例卵巢癌患者、13例女性妇科良性疾病患者及13例同期健康体检女性EDTA抗凝外周全血,采用流式细胞仪分别检测3组单核细胞、CD4+T淋巴细胞、CD8+T淋巴细胞、B淋巴细胞中TLR1、TLR2和TLR6表达水平;TLR1、TLR2和TLR6相应配体(Pam3CSK4、HKLM、FSL-1)分别刺激3组PBMC,采用real-time PCR检测PBMC中前炎症细胞因子IL-1β、IL-6、IL-8、TNF-αm RNA水平。结果:3组细胞亚群中TLR1、TLR2和TLR6均有表达,且主要表达于单核细胞;卵巢癌组单核细胞中TLR1、TLR2、TLR6的表达率分别为69.13%、59.43%、52.99%,显著高于良性疾病组(34.34%、25.32%、15.21%)和健康对照组(36.31%、26.63%、16.43%),差异具有统计学意义(P<0.05);良性疾病组与健康对照组间无显著性差异;3组间CD4+T淋巴细胞、CD8+T淋巴细胞及B淋巴细胞中TLR1、TLR2、TLR6的表达率无显著性差异;3组经TLR2的配体HKLM刺激24 h后,卵巢癌组IL-1βm RNA表达水平显著高于良性疾病组和健康对照组(F=2.05,P<0.05;F=2.19,P<0.05),良性疾病组与健康对照组无显著差异;卵巢癌组IL-6m RNA表达水平显著高于良性疾病组和健康对照组(F=1.40,P<0.05;F=1.99,P<0.05),良性疾病组与健康对照组无显著差异;3组间IL-8及TNF-αm RNA表达水平无显著性差异。3组经TLR6配体FSL-1刺激24 h后,卵巢癌组IL-6 m RNA表达水平显著高于良性疾病组和健康对照组(F=1.30,P<0.05;F=1.69,P<0.05),良性疾病组与健康对照组间无显著性差异;3组间IL-1β、IL-8及TNF-αm RNA表达水平无显著性差异。TLR1配体Pam3CSK4刺激24 h后,3组间IL-1β、IL-6、IL-8及TNF-αm RNA表达水平无显著差异。结论:卵巢癌患者单核细胞中高表达的TLR1、TLR2、TLR6介导的前炎症细胞因子的上调,在卵巢癌炎症微环境的形成和维持中发挥重要作用。
Objective：To explore the pathogenesis of ovarian cancer by investigating the expression of Toll-like receptor 1（TLR1）,TLR2 and TLR6 in patients with ovarian cancer. Methods：We collected peripheral blood mononuclear cells（PBMC） from 13 ovarian cancer patients,13 benign diseases controls,and 13 healthy normal controls. The expression levels of TLR1,TLR2 and TLR6 were measured by flow cytometry in monocytes,CD4＋T lymphocytes,CD8＋T lymphocytes and B lymphocytes. Quantitative real-time PCR was used to measure IL-1β,IL-6,IL-8,and TNF-α in the collected from cells treated with TLR1,TLR2 or TLR6 ligands. Results：TLR1,TLR2 and TLR6 were all expressed in PBMC and they were elevated in monocytes from all three groups. The expression of TLR1,TLR2 and TLR6 from monocytes were higher（P〈 0.05） in ovarian cancer patients（69.13%,59.43%,52.99%,respectively） as compared with benign disease controls（34.34%,25.32%,15.21%,respectively） and healthy normal controls（36.31%,26.63%,16.43%,respectively）,there was no difference between benign disease controls and the healthy normal controls. While between CD4＋T cells,CD8＋T cells,as well as among B cells,no significant difference was found. In concordance with the above results,there was an observable increased expression of inflammatory cytokine interleukin interleukin-1β（IL-1β） upon stimulated by HKLM（TLR2 ligand） in ovarian cancer patients compared to benign disease controls and healthy normal controls（F = 2.05,P〈 0.05;F = 2.19,P〈 0.05）,while no significant difference was noticed between benign disease controls and healthy normal controls;IL-6 was also up-regulated in ovarian cancer patients（F = 1.40,P〈 0.05;F= 1.99,P〈 0.05） when compared with benign disease controls and healthy normal controls,while no significant difference was obvioused between benign disease controls and healthy normal controls;Furthermore,there were differences in IL-8,TNF-α among three groups. Conditioned medium with FSL-1（TLR6 ligand） induced higher amounts of IL-6 expression in ovarian cancer patients than benign disease controls and healthy normal controls（F = 1.30,P〈 0.05;F = 1.69,P〈 0.05）,while no difference was found between benign disease controls and healthy normal controls. In particular,all three groups did not show significant up-regulation of IL-1β,IL-8 or TNF-α production in response to FSL-1 stimulation. Moreover,stimulated with Pam3CSK4（TLR1 / 2 ligand）,IL-1β,IL-6,IL-8 or TNF-α didn＇t show significant up-regulation among all three groups. Conclusion：TLR1,TLR2 and TLR6 were highly expressed in monocytes from ovarian cancer patients,and inflammatory reaction mediated by IL-1β,IL-6 may play an important role in driving ovarian tumor progression.
出处
《南京医科大学学报（自然科学版）》
CAS
CSCD
北大核心
2014年第12期1638-1643,共6页
Journal of Nanjing Medical University(Natural Sciences)
基金
国家自然科学基金(30901344
81272324)
江苏省实验诊断学重点实验室基金(XK 201114)
