Abstract
BACKGROUND: An important aspect of the innate immune response to pathogens is the production of anti-microbial peptides such as cathelicidin-related antimicrobial peptide (CRAMP), the murine homologue of human cathelicidin LL-37. In this study, mechanisms regulating LPS-induction of CRAMP gene expression in mast cells were investigated. NF-kappaB and MAPK pathways were the focus of investigation. METHODS: Mouse bone marrow-derived mast cells were grown in culture and stimulated with LPS. MAPKs and NF-kappaB were monitored by immunoblot analysis. ERK, JNK and p38 MAPK were inhibited using siRNAs or a pharmacological inhibitor. Accumulation of the p65 component of NF-kappaB was inhibited by siRNA and NF-kappaB activation was inhibited by overexpression of I kappaB alpha. MEKK2 or MEKK3 were overexpressed by transfection. The effects of all of these treatments on CRAMP gene expression were monitored by RT-PCR. RESULTS: Inhibition of ERK, JNK or p38 MAPK had little discernible effect on LPS-inducible CRAMP gene expression. Overexpression of MEKK2 or MEKK3 likewise had little impact. However, inhibition of the accumulation of p65 NF-kappaB prevented LPS-induced CRAMP mRNA. An important role for NF-kappaB in CRAMP gene expression was confirmed by overexpression of I kappaB alpha, which reduced both basal and induced levels of CRAMP mRNA. CONCLUSIONS: NF-kappaB, but not MAPKs, plays an important role in LPS-mediated induction of CRAMP gene in mast cells. Defects which inhibit NF-kappaB activity may increase susceptibility to bacterial and viral pathogens which are sensitive to cathelicidins.