The Role Of TLR7Activation On The Proliferation And Migration Of HepG2Cells

Objective To investigate the effect of Toll-like receptor7(TLR7) ligand Gardiquimodon the proliferation and migration of HepG2cells, and to analyze their activiatedsignal transduction pathways.Method HepG2cells were treated with Gardiquimod for different time or differentconcentration,MTT methods were used to analyze the the effect of Gardiquimod onthe proliferation of HepG2cells. FACS methods were used to analyze the effect ofGardiquimod on the proliferation of HepG2cells which were treated with Gardiquimod(2μg/ml)for1-5days. HepG2cells were treated with Gardiquimod (2μg/ml) fordifferent time, total RNA and protein was extracted. The mRNA expression of VEGFand TIMP1were measured by Real-time PCR, and Western blot was used to detect theexpression of Cyclin B1, Cyclin E and the MAPK-ERK1/2signal transduction pathway.And used the specific inhibitor of ERK1/2(PD98059) to block corresponding signalingpathways, then the correlation are analyzed. Wound healing assay were used to detectthe effect of Gardiquimod on migration of HepG2cells.Results TLR7was expressed in HepG2cells, but less than that in peripheral bloodmononuclear cells(PBMC). The results of MTT indicated that Gardiquimod of differenttime or different concentration could suppress the proliferation of HepG2cells. Theresults of FACS showed that after HepG2cells which were treated with Gardiquimod (2μg/ml)for1-5days, the percentage of S phase cells was decline. After the HepG2cellswere treated with Gardiquimod for10min, the mRNA of VEGF in the cells wereobviously reduced. However, when the cells were treated with Gardiquimod for30min,the mRNA of TIMP1in HepG2cells were obviously increased. Western blot results indicated that Gardiquimod (2μg/ml) promoted the expression of Cyclin B1and CyclinE. In addition, the phosphorylation of ERK1/2in the cells are obviously down-regulatedand be associated with the decline of VEGF. What，s more, PD98059can effectivelysuppress the phosphorylation of ERK1/2in HepG2cells. Wound healing assayindicated that after HepG2cells were treated with Gardiquimod (2μg/ml) for differenttime, the migration rate and degree of the cells is obvious reduced.Conclusion TLR7agonist Gardiquimod can suppress the proliferation and migration ofHepG2cells.TLR7activation downergulated the expression of VEGF in HepG2cells,and associated with ERK1/2signal pathway.