Lipoteichoic acid stimulates the proliferation, migration and cytokine production of adult dental pulp stem cells without affecting osteogenic differentiation.

AIM: To model in vitro the contact between adult dental pulp stem cells (DPSCs) and lipoteichoic acid (LTA), a cell wall component expressed at the surface of most Gram-positive bacteria. METHODOLOGY: Human DPSCs obtained from impacted third molars were cultured and exposed to various concentrations of S. aureus LTA (0.1, 1.0 and 10 µg mL-1 ). The effects of LTA on DPSCs proliferation and apoptosis were investigated by MTT assay and flow cytometry. Mineralization of DPSCs was evaluated by alizarin red staining assay. Migration was investigated by microphotographs of wound-healing and Transwell migration assays. Reverse transcription polymerase chain reaction was used to examine the effects of LTA on p65 NF-κB translocation and TLR1, TLR2 or TLR6 regulation. Enzyme-linked immunosorbent assay was used to investigate LTA-stimulated DPSCs cytokine production. One-way or two-way ANOVA and Tukey post hoc multiple comparison were used for statistical analysis. RESULTS: DPSCs expressed TLR1, TLR2 and TLR6 involved in the recognition of various forms of LTA or lipoproteins. Exposure to LTA did not up- or down-regulate the mRNAs of TLR1, TLR2 or TLR6 whilst LPS acted as a potent inducer of them [TLR1 (P ≤ 0.05), TLR2 (P ≤ 0.001) and TLR6 (P ≤ 0.001)]. Translocation of p65 NF-κB to the nucleus was detected in LTA-stimulated cells, but to a lesser extent than LPS-stimulated DPSCs (P ≤ 0.001). The viability of cells exposed to LTA was greater than unstimulated cells, which was attributed to an increased proliferation and not to less cell death [LTA 1 µg mL-1 (P ≤ 0.001) and 10 µg mL-1 (P ≤ 0.01)]. For specific doses of LTA (1.0 µg mL-1 ), adhesion of DPSCs to collagen matrix was disturbed (P ≤ 0.05) and cells enhanced their horizontal mobility (P ≤ 0.001). LTA-stimulated DPSCs released IL-6 and IL-8 in a dose-dependent manner (P ≤ 0.0001). At all concentrations investigated, LTA did not influence osteogenic/odontoblastic differentiation. CONCLUSIONS: Human DPSCs were able to sense the wall components of Gram-positive bacteria likely through TLR2 signalling. Consequently, cells modestly proliferated, increased their migratory behaviour and contributed significantly to the local inflammatory response through cytokine release.

Modulatory role of tachykinin NK1 receptor in cholinergic contraction of mouse trachea.

The role of the NK1 receptor in airway contraction induced by electrical field stimulation (EFS) was evaluated by comparing the response in NK1 receptor knockout mice (NK1R-/-) with that of NK1 receptor wild-type controls (WT). A frequency/response curve on tracheas from NK1R-/- mice and NK1R WT littermates was constructed. After incubation with [3H]choline, [3H]acetylcholine release upon EFS was measured by high-performance liquid chromatography and liquid scintillation counting. The effects of atropine (1 x 10(-6) M), tetrodotoxin (1 x 10(-6) M) and a specific NK1R antagonist (SR140333, 1 x 10(-8) M) were studied, as well as the effects of substance P (1 x 10(-5) M) on precontracted tracheas. Upon EFS, NK1R-/- mice had a significant lower trachea contractility than the NK1R WT animals, accompanied with less [3H]acetylcholine release. Pretreatment with atropine or tetrodotoxin abolished the EFS-induced contraction in both strains. Pretreatment with the NK1R antagonist SR140333 significantly reduced the contractility in the NK1R WT but not in the NK1R-/- mice. Substance P caused a small contraction in both NK1R WT and NK1R-/- mice. Substance P induced a relaxation in precontracted tracheas in NK1R WT but not in NK1R-/- mice. The data presented here provide direct evidence that the NK1 receptor augments cholinergic neurotransmission in mouse trachea.

Airway inflammation and tachykinins: prospects for the development of tachykinin receptor antagonists.

The tachykinins substance P and neurokinin A are contained within sensory airway nerves. Immune cells form an additional source of tachykinins in inflamed airways. Elevated levels of tachykinins have been recovered from the airways of patients with asthma and chronic obstructive pulmonary disease. Airway inflammation leads to an upregulation of tachykinin NK(1) and NK(2) receptors. Preclinical studies have indicated a role for the tachykinin NK(1), NK(2) and NK(3) receptors in bronchoconstriction, airway hyperresponsiveness and airway inflammation caused by allergic and nonallergic stimuli. Compounds that are able to block two or three tachykinin receptors hold promise for the treatment of airways diseases such as asthma and/or chronic obstructive pulmonary disease.