低氧促进口腔鳞癌细胞TLR3及TLR4表达的相关研究

›› 2015, Vol. 35 ›› Issue (10): 806-809.

低氧促进口腔鳞癌细胞TLR3及TLR4表达的相关研究

江琳琳¹,韩伟²,王志勇²,张全安¹,郑勤¹

1. 南京市第二医院
2. 南京市口腔医院

收稿日期:2015-04-27 修回日期:2015-06-03 出版日期:2015-10-28 发布日期:2015-10-27
通讯作者: 郑勤 E-mail:rabbitjiang@sina.com
基金资助:
口腔鳞癌微环境内TLRs/NF-kB/HIF的“交互对话”对其进展的作用机制研究;TLRs/NF-kB与HIF信号相互作用促进口腔鳞癌侵袭转移的机制研究

Study on the expression of TLR3 and TLR4 with the stimulation of hypoxia in OSCC cell lines

Received:2015-04-27 Revised:2015-06-03 Online:2015-10-28 Published:2015-10-27

摘要/Abstract

摘要： 目的　　检测常氧培养条件下口腔鳞癌细胞HSC3中Toll样受体3、4（Toll like receptor 3，4，TLR3、TLR4）的表达水平，并进一步探讨低氧微环境对其表达可能的影响。方法　常氧条件下，将口腔鳞癌细胞HSC3培养至指数增长期，收集细胞，采用Real-time PCR方法在mRNA水平检测TLR3、4的表达，继之，采用Western blotting在蛋白水平检测其表达。模拟体内低氧微环境，在体外采用1% O2浓度分别刺激细胞0、3、6、12及24 h，采用Real-time PCR方法在mRNA水平检测TLR3、4的表达变化，进一步通过Western blotting在蛋白水平检测其表达改变。结果　在常氧培养条件下， HSC3细胞中可检测到TLR3和TLR4的表达。采用低氧刺激可以显著促进TLR3、TLR4的表达水平，其中低氧刺激24 h后，与对照组相比，实验组TLR3的蛋白表达水平增高至(6.2±0.1)倍，而TLR4在低氧刺激6 h后蛋白表达水平可增高至(5.6±0.1)倍，其结果在统计学上具有显著性差异（P＜0.05）。结论　口腔鳞癌细胞HSC3在常氧培养下可检测到TLR3、TLR4的表达，肿瘤低氧微环境可进一步促进其表达水平。

关键词: 口腔鳞状细胞癌, Toll样受体3、4, 低氧

Abstract: Objective　To investigate the expression of Toll like receptor 3,4 (TLR3,4) in oral squamous cell carcinoma (OSCC) cells lines cultured in normoxia, and to further investigate the effect of tumor hypoxia microenvironment on its expression. Methods: OSCC cell line HSC3 was cultured under normoxia conditions. Afterreachingthe exponential growth phase, the cells were collected and Real-time PCR and Western blotting were applied to detect the expression of TLRs at mRNA and protein levels respectively. Tumor hypoxia microenvironment was mimiced by exposing the cells to hypoxia(1%O2) for 0,3,6, 12 and 24h . Both Real-time PCR and Western blotting were applied to detect the expression change of TLR3 and TLR4. Results The expression of TLR3 and TLR4 could be detected in normoxia state. After the stimulation of hypoxia, the expression level improved significantly. Among these, the protein expression of TLR3 could increase to 6.2±0.1 times after the stimulation of hypoxia for 24 hours, while the expression of TLR4 could reach 5.6±0.1 times, which was statistically significant（p＜0.05). Conclusions The expression of TLR3 and TLR4 in HSC3could be detected in normoxia state. Hypoxia could further promote its expression.

Key words: Oral squamous carcinoma cells, Toll like receptor 3,4, Hypoxia

中图分类号:

R739.85

江琳琳韩伟王志勇张全安郑勤. 低氧促进口腔鳞癌细胞TLR3及TLR4表达的相关研究[J]. , 2015, 35(10): 806-809.

参考文献

[1]Nomura N,Miyajima N,Sazuka T,et al.Prediction of the coding sequences of unidentified human genesI. The coding sequences of 40 new genes (KIAA0001-KIAA0040) deduced by analysis of randomly sampled cDNA clones from human immature myeloid cell line KG-1[J].. DNA Res,1994,1(1):27-35
[2]He Z,Huang X,Ni Y,et al.Functional toll-like receptor 3 expressed by oral squamous cell carcinoma induced cell apoptosis and decreasedmigration[J].Oral surgery oral med oral pathol oral radiol,2014,118(1):92-100
[3]Zheng Y1,Ni Y,Huang X,et al.Overexpression of HIF -1α indicates a poor prognosis in tongue carcinoma and may be associated with tumour metastasis[J].Oncol Lett,2013,5(4):1285-1289
[4]Paleja B,Anand A,Chaukar D,et al.Decreased functional response to Toll like receptor ligands in patients with oral cancer[J].Hum Immunol,2013,74(8):927-936
[5]Chuang HC,Huang CC,Chien CY,et al.Toll-like receptor 3-mediated tumor invasion in head and neck cancer[J].Oral Oncol,2012,48(3):226-232
[6]兰卫东，洪兵.Toll样受体4在大鼠炎症牙髓表达的免疫组化研究[J].口腔医学,2013,33(10):672-674
[7]Jouhi L,Renkonen S,Atula T,et al.Different Toll-Like Receptor Expression Patterns in Progression toward Cancer[J].Front Immunol,2014,15;5:638-
[8]Li TT,Ogino S,Qian ZR,et al.Toll-like receptor signaling in colorectal cancer: carcinogenesis to cancer therapy[J].World J Gastroenterol,2014,20(47):17699-708
[9]Pimentel-Nunes P,Gon?alves N,Boal-Carvalho I,et al.Decreased Toll-interacting protein and peroxisome proliferator-activated receptor γ are associated with increased expression of Toll-like receptors in colon carcinogenesis[J].J Clin Pathol,2012,65(4):302-8
[10]Tanaka H,Imaizumi T.Inflammatory chemokine expression via Toll-like receptor 3 signaling in normal human mesangial cells[J].Clin Dev Immunol,2013,2013:984708-
[11]Conroy H,Matshall NA,Mills KH.TLR ligand suppression or enhancement of Treg cells? A double edged sword in immunity to tumors[J].Oncogene,2008,27(2):168-180
[12]Paulos CM,Kaiser A,Wrzesinski C,et al.Toll like receptors in tumor immunotherapy[J].Clin Cancer Res,2007,13(18):5280-5289
[13]罗清琼，陈万涛，胡水清，等.口腔鳞状细胞癌细胞TLR3表达及其配体poly(I：C)的诱导凋亡作用[J].上海交通大学学报（医学版,2011,31(1):5-8
[14]孙祖俊，罗清琼，陈福祥.TLR4介导口腔鳞癌细胞分泌IL6、IL8及VEGF[J].现代免疫学,2010,30(5):395-398
[15]韩生伟,韩伟,泥艳红,等.氯化钴诱导建立口腔鳞状细胞癌低氧模型及其生物学行为初探[J].中华口腔医学杂志,2015,50(3):173-177
[16]Zhou Z,Zhu X,Chen J,et al.The interaction between Toll like receptor 4 signaling pathway and hypoxia inducible factor 1α in lung ischemia reperfusion injury[J].Surrical Research,2014,188(1):290-297
[17]Richa Tewari,Sauray Roy Choudhury,Sadashib Ghosh,et al.Involvement of TNFα induced TLR4-NF-KB and TLR4-HIF-1α feed forward loops in the regulation of inflammatory responses in glioma[J].J Mol Med Berl,2012,90(1):67-80
[18]韩生伟,韩伟,胡勤刚.炎症低氧与口腔肿瘤的发生发展[J].口腔医学研究,2014,30(10):1007-1009