TLR3 activation stimulates cytokine secretion without altering agonist-induced human small airway contraction or relaxation

Cooper PR, Lamb R, Day ND, Branigan PJ, Kajekar R, San Mateo L, Hornby PJ, Panettieri RA Jr. TLR3 activation stimulates cytokine secretion without altering agonist-induced human small airway contraction or relaxation. Am J Physiol Lung Cell Mol Physiol 297:L530-L537, 2009. First published June 19, 2009; doi:10.1152/ajplung.00133.2009.-Respiratory infections exacerbate chronic lung diseases promoting airway inflammation and hyper-reactivity. Toll-like receptor 3 (TLR3) recognizes viral double-stranded (ds) RNA such as polyinosinic-polycytidylic acid [poly(I:C)] and stimulates innate immune responses. The objective of this study was to test the hypothesis that dsRNA promotes lung inflammation and alters airway responsiveness to cholinergic and beta-adrenergic receptor agonists in human lung slices. Human airway smooth muscle (ASM) was incubated for 24 h in poly(I:C) +/- TNF alpha and a TLR3 monoclonal antibody. Precision-cut lung slices (PCLS; 250-mu m thickness) from healthy human lungs containing a small airway were incubated in 0, 10, or 100 mu g/ml poly(I:C) for 24 h. Intravital microscopy of lung slices was used to quantify contractile and relaxation responsiveness to carbachol and isoproterenol, respectively. Supernatants of ASM and PCLS were analyzed for cytokine secretion using a 25-multiplex bead assay. In human ASM, poly(I:C) (0.5 mu g/ml) increased macrophage inflammatory protein-1 alpha (MIP-1 alpha) and RANTES that was prevented by a TLR3 monoclonal receptor antibody. Incubation of human PCLS with poly(I:C) (10 and 100 mu g/ml) had little effect on the log EC50 or maximum drug effect (E-max) for contraction and relaxation in response to carbachol and isoproterenol, respectively. The responsiveness of the same human PCLS to poly(I:C) incubation was confirmed by the robust increase in chemokines and cytokines. In separate experiments, incubation of PCLS with IL-13 or TNF alpha (100 ng/ml) increased airway sensitivity to carbachol. Poly(I:C) promotes inflammatory mediator release that was not associated with enhanced bronchoconstriction or attenuated bronchodilation in normal healthy human lung slices. Transduction at the TLR3 initiated by dsRNA stimulates downstream innate immune responses.