Toll-like receptor-4 agonist inhibits motility and invasion of hepatoblastoma HepG2 cells in vitro.
BACKGROUND
Expression of toll-like receptor-4 (TLR4) on tumor cells is known to mediate innate immune responses that influence tumor cell growth and migration. This study aimed to characterize TLR4 expression and elucidate its functional significance in human hepatoblastoma (HB) cells.


METHODS
Immunohistochemistry (IHC) was used to determine TLR4 expression level and its distribution pattern in HB liver tissues. Transcripts of tumor necrosis factor (TNF)-α, interleukin (IL)-8, matrix metalloproteinase (MMP)-2, MMP-13, tissue inhibitor of metalloproteinases (TIMP)-1, and TIMP-2 in HB HepG2 cells with lipopolysacharide (LPS) treatment were measured by quantitative PCR. Soluble cytokines and peptides in conditioned media were measured by ELISA. MMP-2 activity was determined by using gelatin zymography. Cell motility and invasiveness was determined using wound healing migration and Matrigel invasion assays, respectively.


RESULTS
TLR4 IHC staining demonstrated that TLR4 overexpression in HB liver tissues dramatically vanished after chemotherapy. In vitro study using an HB cell line, HepG2, showed that TLR4 agonist, LPS, significantly decreased transcripts of IL-8 and TNF-α, but did not affect MMP-13 mRNA level. By contrast, LPS only down-regulated IL-8 production and MMP-2 gelatinolytic activity. The latter might be in part due to the increased levels of MMP-2/TIMP-2 complex in conditioned media, thus leading to the decreased motility and invasiveness of HepG2 cells.


CONCLUSIONS
HB cells overexpress TLR4, whereas TLR4 agonistic treatment inhibits migration and invasion of HB HepG2 cells. These findings suggest that TLR4 signaling pathway is a potential therapeutic target for control of HB tumor progression.
Division of Hematology/Oncology, Department of Pediatrics, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.
Chih-Cheng
CC
Hsiao
Ying-Hsien
YH
Kao
Shun-Chen
SC
Huang
Jiin-Haur
JH
Chuang
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use; D002465:Cell Movement / Q000187:drug effects*; D016207:Cytokines / Q000032:analysis / Q000096:biosynthesis; D004797:Enzyme-Linked Immunosorbent Assay; D056945:Hep G2 Cells; D018197:Hepatoblastoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D007150:Immunohistochemistry; D008070:Lipopolysaccharides / Q000494:pharmacology; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D009361:Neoplasm Invasiveness / Q000517:prevention & control*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000819:agonists* / Q000378:metabolism
D016207:Cytokines; D008070:Lipopolysaccharides; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4