title: Gene silencing of Toll-like receptor 2 inhibits proliferation of human liver cancer cells and secretion of inflammatory cytokines.
abstract: BACKGROUND
  Toll-like receptors (TLRs) are key factors in the innate immune system and initiate the inflammatory response to foreign pathogens such as bacteria, fungi and viruses. In the microenvironment of tumorigenesis, TLRs can promote inflammation and cell survival. Toll-like receptor 2/6 (TLR2/6) signaling in tumor cells is regarded as one of the mechanisms of chronic inflammation but it can also mediate tumor cell immune escape and tumor progression. However, the expression of TLR2 and its biological function in the development and progression of hepatocarcinoma have not been investigated. This study aimed to determine the expression of TLRs 1-10 in the established human hepatocellular carcinoma cell line BLE-7402, to investigate the biological effect of TLR2 on cell growth and survival.
  
  
  METHODS
  TLR expression in BLE-7402 cells was assayed by RT-PCR, real-time PCR and flow cytometry (FCM). To further investigate the function of TLR2 in hepatocarcinoma growth, BLE-7402 cells were transfected with recombinant plasmids expressing one of three forms of TLR2 siRNA (sh-TLR2 RNAi(A, B and C)). TLR2 knockdown was confirmed using RT-PCR, real-time PCR and fluorescence microscopy. Tumor cell proliferation was monitored by MTT assay and secreted cytokines in the supernatant of transfected cells were measured by bead-based FCM, the function of TLR2 siRNA was also investigated in vivo.
  
  
  RESULTS
  The BLE-7402 cell line expressed TLRs 2 to 10 at both mRNA and protein levels. TLR2 was the most highly expressed TLR. While all the three siRNAs inhibited TLR2 mRNA and protein expression, sh-TLR2 RNAi(B) had the strongest knockdown effect. TLR2 knockdown with sh-TLR2 RNAi(B) reduced cell proliferation. Furthermore, secretion of IL-6 and IL-8 was also reduced. The result showed a drastic reduction in tumor volume in mice treated with sh-TLR2 RNAi(B).
  
  
  CONCLUSIONS
  These results suggest that TLR2 knockdown inhibit proliferation of cultured hepatocarcinoma cells and decrease the secretion of cytokines. It is suggested that TLR2 silencing may worth further investigations for siRNA based gene therapy in treatment of hepatocarcinoma.
authors: Key Laboratory on Technology for Parasitic Disease Prevention and Control, Ministry of Health, Jiangsu Institute of Parasitic Diseases, Wuxi, Jiangsu, China.
authors: Yuzheng
authors: Y
authors: Huang
authors: Bing
authors: B
authors: Cai
authors: Ming
authors: M
authors: Xu
authors: Zhiqin
authors: Z
authors: Qiu
authors: Yonghui
authors: Y
authors: Tao
authors: Ying
authors: Y
authors: Zhang
authors: Jie
authors: J
authors: Wang
authors: Yongliang
authors: Y
authors: Xu
authors: Yonghua
authors: Y
authors: Zhou
authors: Jing
authors: J
authors: Yang
authors: Xiaofeng
authors: X
authors: Han
authors: Qi
authors: Q
authors: Gao
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics; D016207:Cytokines / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006801:Humans; D007249:Inflammation / Q000378:metabolism; D008113:Liver Neoplasms / Q000473:pathology*; D008297:Male; D051379:Mice; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000235:genetics; D051195:Toll-Like Receptor 2 / Q000172:deficiency* / Q000235:genetics*; D014162:Transfection
chemical_list: D016207:Cytokines; D034741:RNA, Small Interfering; D051195:Toll-Like Receptor 2