Lactobacillus-mediated priming of the respiratory mucosa protects against lethal pneumovirus infection.

The inflammatory response to respiratory virus infection can be complex and refractory to standard therapy. Lactobacilli, when targeted to the respiratory epithelium, are highly effective at suppressing virus-induced inflammation and protecting against lethal disease. Specifically, wild-type mice primed via intranasal inoculation with live or heat-inactivated Lactobacillus plantarum or Lactobacillus reuteri were completely protected against lethal infection with the virulent rodent pathogen, pneumonia virus of mice; significant protection (60% survival) persisted for at least 13 wk. Protection was not unique to Lactobacillus species, and it was also observed in response to priming with nonpathogenic Gram-positive Listeria innocua. Priming with live lactobacilli resulted in diminished granulocyte recruitment, diminished expression of multiple proinflammatory cytokines (CXCL10, CXCL1, CCL2, and TNF), and reduced virus recovery, although we have demonstrated clearly that absolute virus titer does not predict clinical outcome. Lactobacillus priming also resulted in prolonged survival and protection against the lethal sequelae of pneumonia virus of mice infection in MyD88 gene-deleted (MyD88(-/-)) mice, suggesting that the protective mechanisms may be TLR-independent. Most intriguing, virus recovery and cytokine expression patterns in Lactobacillus-primed MyD88(-/-) mice were indistinguishable from those observed in control-primed MyD88(-/-) counterparts. In summary, we have identified and characterized an effective Lactobacillus-mediated innate immune shield, which may ultimately serve as critical and long-term protection against infection in the absence of specific antiviral vaccines.

Prolonged exposure to NT-3 attenuates cholinergic nerve-mediated contractions in cultured murine airways.

Chronic airway inflammation may induce subsequent airway hyper-responsiveness (AHR) including pathological alteration of neural activity. Asthmatic airways contain elevated levels of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) albeit, their effect on neural activity is unclear. This study evaluates the effects of NT-3 and BDNF on nerve mediated airway contractions in vitro. Tracheal segments from BALB/c J mice were cultured for 4 days with NT-3 or BDNF. Responsiveness to electric field stimulation (EFS) was evaluated in organ-bath and innervation patterns were examined by quantitative immunohistochemistry. In cultured segments the EFS-induced contractions were inhibited by tetrodotoxin or atropine. NT-3 reduced the EFS contractions in a concentration-dependent manner whereas BDNF had no effect. The amount of nerve fibers, found in conjunction with the tracheal smooth muscle, was similar in NT-3 treated and control segments. In conclusion, NT-3 attenuates cholinergic nerve-mediated contractions of airway in vitro. Considering the elevated levels of NT-3 found in asthmatic airways, the findings imply a protective role of NT-3 in AHR.

TNF alpha reduces tachykinin, PGE2-dependent, relaxation of the cultured mouse trachea by increasing the activity of COX-2.

1. Chronic inflammation is a central feature of asthma. The inflammatory cytokine tumour necrosis factor alpha (TNFalpha) has been implicated in this disease, and is known to alter airway smooth muscle functionally. 2. The aim of this study was to investigate the influence of TNFalpha on tachykinin-induced airway relaxation. Mouse tracheae were cultured in the absence and presence of TNFalpha for 1 or 4 days. 3. In the absence of TNFalpha, substance P (SP) and neurokinin A (NKA) induced comparable levels of relaxation in fresh and cultured segments. Functional studies with selective antagonists/inhibitors indicated that the relaxation was mediated by the NK(1) receptor coupled to cyclooxygenase (COX)-2 activation and subsequent release of prostaglandin E(2) (PGE(2)). TNFalpha attenuated SP- and NKA-induced relaxation in a time- and concentration-dependent manner, decreasing the ability of PGE(2) to relax tissues. 4. Further studies indicated that TNFalpha elevated COX-2 activity and that concomitant inhibition of COX-2 reversed TNFalpha-attenuated PGE(2) relaxation. Culture with PGE(2) decreased SP- and PGE(2)-mediated relaxation, further implicating the activity of COX-2 in the attenuation of tachykinin signalling. 5. Gene expression analysis demonstrated that TNFalpha increased the expression of smooth muscle COX-2, PGE(2) synthase and EP(2) receptor mRNA, and decreased the expression of the EP(4) receptor. 6. Overall, these results show that NK(1) receptor-mediated relaxation induced by PGE(2) is attenuated by prolonged TNFalpha stimulation. Increased COX-2 activity induced by TNFalpha appears to be central to this process.

Toll-like receptor stimulation induces airway hyper-responsiveness to bradykinin, an effect mediated by JNK and NF-kappa B signaling pathways.

Airway infections induce hyper-responsiveness in asthmatic patients. Toll-like receptors (TLR) mediate inflammatory responses to microbes. Occurrence and effects of TLR2, TLR3 and TLR4 were examined in a mouse organ culture model of asthma focusing on the smooth muscle responses to bradykinin. TLR2, TLR3 and TLR4 mRNA, and TLR2 and TLR4 immunoreactivity were detected in the tracheal muscle layer. Tracheal organ culture for 1 or 4 days with lipopolysaccharide (LPS; TLR2/4 agonist) or polyinosinic polycytidylic acid (poly-I-C; TLR3 agonist) enhanced bradykinin- and [des-Arg(9)]-bradykinin-induced contractions. Simultaneous LPS and poly-I-C treatment resulted in synergistic enhancement of bradykinin-induced contraction. In carbachol-pre-contracted segments TLR stimulation induced less potent relaxations to bradykinin and [des-Arg(9)]-bradykinin. The LPS and poly-I-C enhancement of bradykinin-induced contraction was inhibited by the transcriptional inhibitor actinomycin-D, dexamethasone, the proteasome inhibitor MG-132 and the c-Jun N-terminal kinase (JNK) inhibitor SP600125. LPS and poly-I-C induced translocation of NF-kappa B p65 to the nucleus and up-regulation of kinin B(1) and B(2) receptor mRNA. In summary, TLR2, TLR3 and TLR4 are expressed in the mouse tracheal smooth muscle. Costimulation of these receptors results in NF-kappa B- and JNK-mediated transcription of B(1) and B(2) receptor, inducing hyper-responsiveness to bradykinin.