Toll-like receptor 4 signaling promotes epithelial-mesenchymal transition in human hepatocellular carcinoma induced by lipopolysaccharide

BMC Medicine

Li-Xin Wei¹  Meng-Chao Wu¹  Qiu-Dong Zhao¹  Xue Zhao¹  Lu Gao¹  Rong Li¹  Yan Liu¹  Jing Hou¹  Shan-Shan Zhang¹  Kai Sun¹  Zhi-Peng Han¹  Ying-Ying Jing¹ 

[1] Tumor Immunology and Gene Therapy Center, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai, PR China

关键词: Human hepatocellular carcinoma;    Lipopolysaccharide;    Epithelial-mesenchymal transition;    Toll-like receptor 4;   

Others  :  857360
DOI  :  10.1186/1741-7015-10-98

 received in 2012-02-15, accepted in 2012-08-31,  发布年份 2012

【 摘 要 】

Background

The endotoxin level in the portal and peripheral veins of hepatocellular carcinoma (HCC) patients is higher and lipopolysaccharide (LPS), a cell wall constituent of gram-negative bacteria, has been reported to inhibit tumor growth. However, in this study, we found that LPS-induced toll-like receptor 4 (TLR4) signaling was involved in tumor invasion and survival, and the molecular mechanism was investigated,

Methods

Four HCC cell lines and a splenic vein metastasis of the nude mouse model were used to study the invasion ability of LPS-induced HCC cells and the epithelia-mesenchymal transition (EMT) in vitro and in vivo. A total of 106 clinical samples from HCC patients were used to evaluate TLR4 expression and analyze its association with clinicopathological characteristics

Results

The in vitro and in vivo experiments demonstrated that LPS could significantly enhance the invasive potential and induce EMT in HCC cells with TLR4 dependent. Further studies showed that LPS could directly activate nuclear factor kappa B (NF-κB) signaling through TLR4 in HCC cells. Interestingly, blocking NF-κB signaling significantly inhibited transcription factor Snail expression and thereby inhibited EMT occurrence. High expression of TLR4 in HCC tissues was strongly associated with both poor cancer-free survival and overall survival in patients.

Conclusions

Our results indicate that TLR4 signaling is required for LPS-induced EMT, tumor cell invasion and metastasis, which provide molecular insights for LPS-related pathogenesis and a basis for developing new strategies against metastasis in HCC.

【 授权许可】

   
2012 Jing et al; licensee BioMed Central Ltd.

【 图 表 】

【 参考文献 】

• [1]Cordon-Cardo C, Prives C: At the crossroads of inflammation and tumorigenesis. J Exp Med 1999, 190:1367-1370.
• [2]Nolan JP, Leibowitz AI: Endotoxins in liver disease. Gastroenterology 1978, 75:765-766.
• [3]Triger DR, Boyer TD, Levin J: Portal and systemic bacteraemia and endotoxaemia in liver disease. Gut 1978, 19:935-939.
• [4]Schwabe RF, Seki E, Brenner DA: Toll-like receptor signaling in the liver. Gastroenterology 2006, 130:1886-1900.
• [5]Iwasaki A, Medzhitov R: Toll-like receptor control of the adaptive immune responses. Nat Immunol 2004, 5:987-995.
• [6]Pidgeon GP, Harmey JH, Kay E, Da Costa M, Redmond HP, Bouchier-Hayes DJ: The role of endotoxin/lipopolysaccharide in surgically induced tumour growth in a murine model of metastatic disease. Br J Cancer 1999, 81:1311-1317.
• [7]Harmey JH, Bucana CD, Lu W, Byrne AM, McDonnell S, Lynch C, Bouchier-Hayes D, Dong Z: Lipopolysaccharide-induced metastatic growth is associated with increased angiogenesis, vascular permeability and tumor cell invasion. Int J Cancer 2002, 101(5):415-422.
• [8]Wang JH, Manning BJ, Wu QD, Blankson S, Bouchier-Hayes D, Redmond HP: Endotoxin/lipopolysaccharide activates NF-kappa B and enhances tumor cell adhesion and invasion through a beta 1 integrin-dependent mechanism. J Immunol 2003, 170:795-804.
• [9]Liu X, Liang J, Li G: Lipopolysaccharide promotes adhesion and invasion of hepatoma cell lines HepG2 and HepG2.2.15. Mol Biol Rep 2010, 37:2235-2239.
• [10]Ikebe M, Kitaura Y, Nakamura M, Tanaka H, Yamasaki A, Nagai S, Wada J, Yanai K, Koga K, Sato N, Kubo M, Tanaka M, Onishi H, Katano M: Lipopolysaccharide (LPS) increases the invasive ability of pancreatic cancer cells through the TLR4/MyD88 signaling pathway. J Surg Oncol 2009, 100:725-731.
• [11]Akira S, Takeda K: Toll-like receptor signalling. Nat Rev 2004, 4:499-511.
• [12]Hua D, Liu MY, Cheng ZD, Qin XJ, Zhang HM, Chen Y, Qin GJ, Liang G, Li JN, Han XF, Liu DX: Small interfering RNA-directed targeting of Toll-like receptor 4 inhibits human prostate cancer cell invasion, survival, and tumorigenicity. Mol Immunol 2009, 46:2876-2884.
• [13]Yu LX, Yan HX, Liu Q, Yang W, Wu HP, Dong W, Tang L, Lin Y, He YQ, Zou SS, Wang C, Zhang HL, Cao GW, Wu MC, Wang HY: Endotoxin accumulation prevents carcinogen-induced apoptosis and promotes liver tumorigenesis in rodents. Hepatology 2010, 52:1322-1333.
• [14]Gonzalez-Reyes S, Marin L, Gonzalez L, Gonzalez LO, del Casar JM, Lamelas ML, Gonzalez-Quintana JM, Vizoso FJ: Study of TLR3, TLR4 and TLR9 in breast carcinomas and their association with metastasis. BMC Cancer 2010, 10:665. BioMed Central Full Text
• [15]Xu Z, Ren T, Xiao C, Li H, Wu T: Nickel promotes the invasive potential of human lung cancer cells via TLR4/MyD88 signaling. Toxicology 2011, 285:25-30.
• [16]Wang EL, Qian ZR, Nakasono M, Tanahashi T, Yoshimoto K, Bando Y, Kudo E, Shimada M, Sano T: High expression of Toll-like receptor 4/myeloid differentiation factor 88 signals correlates with poor prognosis in colorectal cancer. Br J Cancer 2010, 102:908-915.
• [17]Larue L, Bellacosa A: Epithelial-mesenchymal transition in development and cancer: role of phosphatidylinositol 3' kinase/AKT pathways. Oncogene 2005, 24:7443-7454.
• [18]Zhao L, Yang R, Cheng L, Wang M, Jiang Y, Wang S: LPS-induced epithelial-mesenchymal transition of intrahepatic biliary epithelial cells. J Surg Res 2011, 171:819-25.
• [19]Wu F, Yang LY, Li YF, Ou DP, Chen DP, Fan C: Novel role for epidermal growth factor-like domain 7 in metastasis of human hepatocellular carcinoma. Hepatology 2009, 50:1839-1850.
• [20]Yang MH, Chen CL, Chau GY, Chiou SH, Su CW, Chou TY, Peng WL, Wu JC: Comprehensive analysis of the independent effect of twist and snail in promoting metastasis of hepatocellular carcinoma. Hepatology 2009, 50:1464-1474.
• [21]Zhang X, Liu S, Hu T, Liu S, He Y, Sun S: Up-regulated microRNA-143 transcribed by nuclear factor kappa B enhances hepatocarcinoma metastasis by repressing fibronectin expression. Hepatology 2009, 50:490-499.
• [22]Sun T, Zhao N, Zhao XL, Gu Q, Zhang SW, Che N, Wang XH, Du J, Liu YX, Sun BC: Expression and functional significance of Twist1 in hepatocellular carcinoma: its role in vasculogenic mimicry. Hepatology 2010, 51:545-556.
• [23]Zhang J, Chen Y, Xin XL, Li QN, Li M, Lin LP, Geng MY, Ding J: Oligomannurarate sulfate blocks tumor growth by inhibiting NF-kappaB activation. Acta Pharmacol Sin 2010, 31:375-381.
• [24]Szajnik M, Szczepanski MJ, Czystowska M, Elishaev E, Mandapathil M, Nowak-Markwitz E, Spaczynski M, Whiteside TL: TLR4 signaling induced by lipopolysaccharide or paclitaxel regulates tumor survival and chemoresistance in ovarian cancer. Oncogene 2009, 28:4353-4363.
• [25]Huber MA, Azoitei N, Baumann B, Grunert S, Sommer A, Pehamberger H, Kraut N, Beug H, Wirth T: NF-kappaB is essential for epithelial-mesenchymal transition and metastasis in a model of breast cancer progression. J Clin Invest 2004, 114:569-581.
• [26]Condeelis J, Segall JE: Intravital imaging of cell movement in tumours. Nat Rev Cancer 2003, 3:921-930.
• [27]Dong R, Wang Q, He XL, Chu YK, Lu JG, Ma QJ: Role of nuclear factor kappa B and reactive oxygen species in the tumor necrosis factor-alpha-induced epithelial-mesenchymal transition of MCF-7 cells. Braz J Med Biol Res 2007, 40:1071-1078.
• [28]Molteni M, Marabella D, Orlandi C, Rossetti C: Melanoma cell lines are responsive in vitro to lipopolysaccharide and express TLR-4. Cancer Lett 2006, 235:75-83.
• [29]van Zijl F, Zulehner G, Petz M, Schneller D, Kornauth C, Hau M, Machat G, Grubinger M, Huber H, W Mikulits: Epithelial to mesenchymal transition in hepatocellular carcinoma. Future Oncol 2009, 5:1169-1179.
• [30]Nakashima H, Nakamura M, Yamaguchi H, Yamanaka N, Akiyoshi T, Koga K, Yamaguchi K, Tsuneyoshi M, Tanaka M, Katano M: Nuclear factor-kappaB contributes to hedgehog signaling pathway activation through sonic hedgehog induction in pancreatic cancer. Cancer Res 2006, 66:7041-7049.
• [31]Kojima M, Morisaki T, Izuhara K, Uchiyama A, Matsunari Y, Katano M, Tanaka M: Lipopolysaccharide increases cyclo-oxygenase-2 expression in a colon carcinoma cell line through nuclear factor-kappa B activation. Oncogene 2000, 19:1225-1231.
• [32]Wu Y, Deng J, Rychahou PG, Qiu S, Evers BM, Zhou BP: Stabilization of snail by NF-kappaB is required for inflammation-induced cell migration and invasion. Cancer Cell 2009, 15:416-428.
• [33]Barbera MJ, Puig I, Dominguez D, Julien-Grille S, Guaita-Esteruelas S, Peiro S, Baulida J, Franci C, Dedhar S, Larue L, García de Herreros A: Regulation of Snail transcription during epithelial to mesenchymal transition of tumor cells. Oncogene 2004, 23:7345-7354.
• [34]Criswell TL, Arteaga CL: Modulation of NFkappaB activity and E-cadherin by the type III transforming growth factor beta receptor regulates cell growth and motility. J Biol Chem 2007, 282:32491-32500.
• [35]Franco DL, Mainez J, Vega S, Sancho P, Murillo MM, de Frutos CA, Del Castillo G, Lopez-Blau C, Fabregat I, Nieto MA: Snail1 suppresses TGF-beta-induced apoptosis and is sufficient to trigger EMT in hepatocytes. J Cell Sci 2010, 123:3467-3477.
• [36]Batlle E, Sancho E, Franci C, Dominguez D, Monfar M, Baulida J, Garcia De Herreros A: The transcription factor snail is a repressor of E-cadherin gene expression in epithelial tumour cells. Nat Cell Biol 2000, 2:84-89.
• [37]Cano A, Perez-Moreno MA, Rodrigo I, Locascio A, Blanco MJ, del Barrio MG, Portillo F, Nieto MA: The transcription factor snail controls epithelial-mesenchymal transitions by repressing E-cadherin expression. Nat Cell Biol 2000, 2:76-83.
• [38]Cammarota R, Bertolini V, Pennesi G, Bucci EO, Gottardi O, Garlanda C, Laghi L, Barberis MC, Sessa F, Noonan DM, Albini A: The tumor microenvironment of colorectal cancer: stromal TLR-4 expression as a potential prognostic marker. J Transl Med 2010, 8:112. BioMed Central Full Text
• [39]Velazquez RF, Rodriguez M, Navascues CA, Linares A, Perez R, Sotorrios NG, Martinez I, Rodrigo L: Prospective analysis of risk factors for hepatocellular carcinoma in patients with liver cirrhosis. Hepatology 2003, 37:520-527.
• [40]Poon RT, Fan ST, Lo CM, Liu CL, Wong J: Difference in tumor invasiveness in cirrhotic patients with hepatocellular carcinoma fulfilling the Milan criteria treated by resection and transplantation: impact on long-term survival. Ann Surg 2007, 245:51-58.