title: Downregulation of toll-like receptor 4 induces suppressive effects on hepatitis B virus-related hepatocellular carcinoma via ERK1/2 signaling.
abstract: BACKGROUND
  Hepatitis B virus (HBV) infection is a major risk factor which can lead to development of hepatocellular carcinoma (HCC). In this study, we aimed to explore the effects of toll-like receptor 4 (TLR4) downregulation on the growth and survival of HBV-related HCC cells and to examine the molecular mechanisms been involved.
  
  
  METHODS
  The expression levels of TLR4 were examined in a panel of HCC cell lines (HepG2, SMMC7721, Huh7, HepG2.2.15 and Hep3B). The effects of TLR4 downregulation on the proliferation, apoptosis, and tumorigenicity of HBV-related HepG2.2.15 cells were determined. The effects of TLR4 downregulation on multiple signaling pathways were also measured. Co-immunoprecipitation and immunofluoresence staining assays were performed to investigate the interaction between TLR4 and HBV X protein (HBx).
  
  
  RESULTS
  The mRNA and protein levels of TLR4 were significantly increased in HepG2.2.15 cells than those in the other cells which have been studied. Downregulation of TLR4 significantly decreased the proliferation and induced G2/M cell cycle arrest and apoptosis in HepG2.2.15 cells. TLR4 depletion inhibited HepG2.2.15 cell colony formation and tumor growth in nude mice. TLR4 silencing decreased the phosphorylation of ERK1/2 but not JNK1/2, p38, or NF-κB. Chemical inhibition of ERK1/2 approximately phenocopied the growth-suppressive effect of TLR4 downregulation on HepG2.2.15 cells. In addition, TLR4 showed a physical interaction with HBx.
  
  
  CONCLUSIONS
  Taken together, TLR4 plays a tumor-promoting role in HBV-related HCC cells, which is associated with regulation of ERK1/2 activation and interaction with HBx. Therefore, TLR4 may be a potential therapeutic target for HBV-related HCC.
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. yitingwang_1111@163.com.
authors.forename: Yiting
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. cjdl123@163.com.
authors.forename: Jing
authors.initials: J
authors.lastname: Cai
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. rememberme0915@126.com.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Zeng
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. yjchen0506@163.com.
authors.forename: Yajie
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. yanweishandian@126.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Yan
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. oyym89@163.com.
authors.forename: Yuming
authors.initials: Y
authors.lastname: Ouyang
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. xd870824@163.com.
authors.forename: Dan
authors.initials: D
authors.lastname: Xiao
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. zzm@163.com.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Zeng
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. huanglong914@gmail.com.
authors.forename: Long
authors.initials: L
authors.lastname: Huang
authors.affiliation: Department of Oncology, the Second Affiliated Hospital of Nanchang University, Nanchang, 330006, Jiangxi Province, China. awliu666@163.com.
authors.forename: Anwen
authors.initials: A
authors.lastname: Liu
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics; D006528:Carcinoma, Hepatocellular / Q000209:etiology* / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D015536:Down-Regulation; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D056945:Hep G2 Cells; D006509:Hepatitis B / Q000150:complications*; D006515:Hepatitis B virus; D064593:Heterografts; D006801:Humans; D008113:Liver Neoplasms / Q000209:etiology* / Q000378:metabolism* / Q000473:pathology; D020935:MAP Kinase Signaling System; D008297:Male; D051379:Mice; D011485:Protein Binding; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism; D047368:Tumor Burden; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D051197:Toll-Like Receptor 4; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein