Aims: In this work we aimed to investigate the expression of TLR9 protein in the murine hepatocyte cell line TIB-73, compared to macrophage-like J774 cells, by Western blot analysis, and the role played by ERK 1/2 MAP kinase in the intracellular signals triggered by stimulation with CpG and non-CpG phosphodiester-ODN, and their more stable phosphorothioate-modified analogues.
Results: TIB-73 hepatocytes express TLR9 protein. CpG and non-CpG ODN stimulation activated ERK 1/2 MAPK signal pathway in both hepatocytes and J774 murine macrophages. As expected, their phosphorothioate-CpG and non-CpG ODN analogues induced higher levels of ERK1/2 phosphorylation in TIB-73 cells, even higher than that induced in J774 cells under the same conditions. Phosphorylation of ERK 1/2 induced by synthetic ODN is dose-response dependent, being maximal at 100 microg/ml. Pretreatment of hepatocytes with an inhibitor of MEK-1 abrogated phosphorylation of ERK1/2 kinase.
Conclusions: TIB-73 hepatocytes constitutively express TLR9 and respond to synthetic ODN stimulation through a high ERK1/2 phosphorylation independent of CpG motifs. Slight differences were found on ERK1/2 activation when using phosphorothioate versus phosphodiester oligonucleotides.