Mycobacterium bovis BCG Toll-like receptors 2 and 4 cooperation increases the innate epithelial immune response.

Background: Although CXCL8 is known to be important both in enhancing local antimycobacterial activity and in driving recruitment of leukocyte subsets during inflammatory processes, little information is known about the relationship between transcriptional upregulation of the Mycobacterium bovis BCG-induced CXCL8 secretion and TLR signaling.

Methods: Phosphorylation of ERK1/2 was analyzed by immunoblot analysis. The levels of immunoreactive CXCL8 were measured with cytokine-specific commercial ELISA kits. By using antibodies against both TLR2 and TLR4, the levels of CXCL8 mRNA were determined by RT-PCR.

Results: In this study, we found that the inhibition of TLR2 significantly, and that of TLR4 partially, decreased the ERK1/2 activation and the subsequent CXCL8 secretion induced by the M. bovis BCG when neutralizing antibodies to TLR2 or TLR4 were applied to human epithelial cells. Moreover, it is important to note that the levels of CXCL8 secretion and mRNA were markedly (94%) reduced by functional blocking of both TLR2 and TLR4. Under the same conditions, control stimulation with TLR4 (LPS) was markedly blocked with an anti-TLR4 antibody. Finally, when ERK1/2 activity was inhibited in TLR2- or TLR4-expressing cells, M. bovis BCG-dependent CXCL8 production resulted in a significant inhibition.

Conclusions: Together these results suggest that the M. bovis BCG, acting through both TLR2 and TLR4, induces the activation of the ERK1/2 MAPK pathway, which in turn plays a major role in CXCL8 secretion.