rhLBP Modulates Expression of Pro-inflammatory Cytokines and hBD-2 in HOKs

Abstract

Objectives: LPS-binding protein (LBP) plays an important role in innate host response to microbial challenge. Lately, it has been shown that LBP could be anchored in the cell surface and a certain role of this intercalated LBP has been proposed. Our recent studies find that LBP is expressed in human gingiva, and its expression is associated with periodontal conditions. The present study investigated the potential effects of LBP on the innate host response in human oral keratinocytes (HOKs). Methods: HOKs were treated with various concentrations of rhLBP (10ng/ml, 100ng/ml and 1µg/ml) for 1 h and 4 h for mRNA detection, and 13 h and 25 h for peptide detection, respectively. For the blocking assay, cells were pre-incubated with anti-TLR4 mAb or anti-TLR2 mAb at 10µg/ml for 1 h prior to incubation with 100ng/ml of rhLBP for 4 h and 25 h. The mRNA expression of IL-6, IL-8, TNF-α and hBD-2 was detected by real-time PCR. The peptides of IL-6, IL-8 and hBD-2 were analyzed by ELISA.

Results: rhLBP could significantly up-regulate IL-6, IL-8 and TNF-α mRNA expression at 4 h, but no up-regulation was observed at 1 h. It up-regulated IL-6 and IL-8 peptide expression dose dependently at 25 h, but no up-regulation was observed at 13 h. On the other side, rhLBP could down-regulate hBD-2 mRNA and peptide expression at 4 h and 13 h, respectively. Blocking assay revealed that rhLBP-induced pro-inflammatory cytokine expression could be partially neutralized by TLR2 antibody, but not by TLR4 antibody.

Conclusions: The present study shows that rhLBP alone could up-regulate IL-6, IL-8, TNF-α and down-regulate hBD-2 expression, and this rhLBP-mediated effect on innate host response may be through TLR2 but not TLR4.