The short-chain free fatty acid receptor FFAR3 is expressed and potentiates contraction in human airway smooth muscle.

Emerging evidence suggests that gut microbiota-derived short-chain fatty acids (SCFAs; acetate, propionate, and butyrate) are important modulators of the inflammatory state in diseases such as asthma. However, the functional expression of the Gi protein-coupled free fatty acid receptors (FFAR2/GPR43 and FFAR3/GPR41) has not been identified on airway smooth muscle (ASM). Classically, acute activation of Gi-coupled receptors inhibits cyclic AMP (cAMP) synthesis, which impairs ASM relaxation and can also induce crosstalk between Gi- and Gq-signaling pathways, potentiating increases in intracellular Ca2+ concentration ([Ca2+]i), favoring ASM contraction. In contrast, chronic activation of Gi-coupled receptors can sensitize adenylyl cyclase resulting in increased cAMP synthesis favoring relaxation. We questioned whether the Gi-coupled FFAR2 or FFAR3 is expressed in human ASM, whether they modulate cAMP and [Ca2+]i, and whether SCFAs modulate human ASM tone. We detected the protein expression of FFAR3 but not FFAR2 in native human ASM and primary cultured human airway smooth muscle (HASM) cells. In HASM cells, acute activation of FFAR3 with SCFAs inhibited forskolin-stimulated cAMP accumulation, but chronic activation did not sensitize cAMP synthesis. SCFAs induced [Ca2+]i increases that were attenuated by pertussis toxin, gallein, U73122, or xestospongin C. Acute treatment with SCFAs potentiated acetylcholine-stimulated [Ca2+]i increases and stress fiber formation in cells and contraction of ex vivo human airway tissues. In contrast, chronic pretreatment of human ASM with propionate did not potentiate airway relaxation. Together, these findings demonstrate that FFAR3 is expressed in human ASM and contributes to ASM contraction via reduced cAMP and increased [Ca2+]i.

The free fatty acid receptor 1 promotes airway smooth muscle cell proliferation through MEK/ERK and PI3K/Akt signaling pathways.

Obesity is a risk factor for asthma and influences airway hyperresponsiveness, which is in part modulated by airway smooth muscle proliferative remodeling. Plasma free fatty acids (FFAs) levels are elevated in obese individuals, and long-chain FFAs act as endogenous ligands for the free fatty acid receptor 1 (FFAR1), which couples to both Gq and Gi proteins. We examined whether stimulation of FFAR1 induces airway smooth muscle cell proliferation through classical MEK/ERK and/or phosphoinositide 3-kinase (PI3K)/Akt signaling pathways. The long-chain FFAs (oleic acid and linoleic acid) and a FFAR1 agonist (GW9508) induced human airway smooth muscle (HASM) cell proliferation, which was inhibited by the MEK inhibitor U0126 and the PI3K inhibitor LY294002 . The long-chain FFAs and GW9508 increased phosphorylation of ERK, Akt, and p70S6K in HASM cells and freshly isolated rat airway smooth muscle. Downregulation of FFAR1 in HASM cells by siRNA significantly attenuated oleic acid-induced phosphorylation of ERK and Akt. Oleic acid-induced ERK phosphorylation was blocked by either the Gαi-protein inhibitor pertussis toxin or U0126 and was partially inhibited by either the Gαq-specific inhibitor YM-254890 or the Gßγ signaling inhibitor gallein. Oleic acid significantly inhibited forskolin-stimulated cAMP activity, which was attenuated by pertussis toxin. Akt phosphorylation was inhibited by pertussis toxin, the ras inhibitor manumycin A, the Src inhibitor PP1, or LY294002 . Phosphorylation of p70S6K by oleic acid or GW9508 was significantly inhibited by LY294002 , U0126, and the mammalian target of rapamycin (mTOR) inhibitor rapamycin. In conclusion, the FFAR1 promoted airway smooth muscle cell proliferation and p70S6K phosphorylation through MEK/ERK and PI3K/Akt signaling pathways.

The dopamine D1 receptor is expressed and induces CREB phosphorylation and MUC5AC expression in human airway epithelium.

Background: Dopamine receptors comprise two subgroups, Gs protein-coupled "D1-like" receptors (D1, D5) and Gicoupled "D2-like" receptors (D2, D3, D4). In airways, both dopamine D1 and D2 receptors are expressed on airway smooth muscle and regulate airway smooth muscle force. However, functional expression of the dopamine D1 receptor has never been identified on airway epithelium. Activation of Gs-coupled receptors stimulate adenylyl cyclase leading to cyclic AMP (cAMP) production, which is known to induce mucus overproduction through the cAMP response element binding protein (CREB) in airway epithelial cells. We questioned whether the dopamine D1 receptor is expressed on airway epithelium, and whether it promotes CREB phosphorylation and MUC5AC expression. Methods: We evaluated the protein expression of the dopamine D1 receptor on native human airway epithelium and three sources of cultured human airway epithelial cells including primary cultured airway epithelial cells, the bronchial epithelial cell line (16HBE14o-), and the pulmonary mucoepidermoid carcinoma cell line (NCI-H292) using immunohistochemistry and immunoblotting. To characterize the stimulation of cAMP through the dopamine D1 receptor, 16HBE14o- cells and NCI-H292 cells were treated with dopamine or the dopamine D1 receptor agonists (SKF38393 or A68930) before cAMP measurements. The phosphorylation of CREB by A68930 in both 16HBE14o- and NCI-H292 cells was measured by immunoblot. The effect of dopamine or A68930 on the expression of MUC5AC mRNA and protein in NCI-H292 cells was evaluated by real-time PCR and immunofluorescence staining, respectively. Results: The dopamine D1 receptor protein was detected in native human airway epithelium and three sources of cultured human airway epithelial cells. Dopamine or the dopamine D1-like receptor agonists stimulated cAMP production in 16HBE14o- cells and NCI-H292 cells, which was reversed by the selective dopamine D1-like receptor antagonists (SCH23390 or SCH39166). A68930 significantly increased phosphorylation of CREB in both 16HBE14o- and NCI-H292 cells, which was attenuated by the inhibitors of PKA (H89) and MEK (U0126). Expression of MUC5AC mRNA and protein were also increased by either dopamine or A68930 in NCI-H292 cells. Conclusions: These results suggest that the activation of the dopamine D1 receptor on human airway epithelium could induce mucus overproduction, which could worsen airway obstructive symptoms.

Efficacy of intrathecal esmolol on heat-evoked responses in a postoperative pain model.

Perioperative tachycardia and hypertension are often treated with esmolol, a short-acting ß1-adrenoceptor antagonist. Besides its cardiac effect, esmolol is reported to exert antinociceptive effects. This study examined the efficacy of intrathecal (IT) esmolol on pain responses in a postoperative pain model. Male Sprague-Dawley rats (250-300 g) were anesthetized with sevoflurane and an IT catheter was implanted. Six days after catheter implantation, a postoperative pain model was established by plantar incision under sevoflurane anesthesia. Withdrawal latencies were assessed by applying a focused radiant heat source before plantar incision; 1 day after the incision (before esmolol administration); and 5, 10, and 15 minutes after bolus administration of IT esmolol. Plantar incision produced hypersensitivity in the postoperative pain model expressed as decreased withdrawal latency to heat stimulation (before incision: 13.9 ± 0.29 seconds and 1 day after incision: 6.3 ± 0.26 seconds). These decreased latencies caused by incision were significantly increased by esmolol administration (40 µg, 80 µg) at 5 minutes (10.7 ± 1.16 seconds, 10.5 ± 1.16 seconds). No postoperative antinociceptive effects of esmolol were observed at 10 or 15 minutes. IT administration of esmolol produced antinociceptive effects of short duration in a rat postoperative pain model. These results suggest that IT esmolol could offer a new strategy for managing perioperative pain, although an alternative approach is necessary to lengthen the duration of the analgesia.

[Comparison of Pentax-AWS, GlideScope Cobalt, and Macintosh laryngoscope in patients for nasotracheal intubation].

BACKGROUND: We compared the time for instrumentation in nasotracheal intubation using Pentax-AWS and Glidescope to that using the Macintosh laryngoscope in patients with normal airway. METHODS: After local ethics board approval, 60 patients requiring dental or oral surgery were allocated randomly to Pentax-AWS, Glidescope and Macintosh groups. One experienced anesthesiologist performed nasotracheal intubation in all patients. RESULTS: The times for instrumentation using Pentax-AWS and Glidescope, and Macintosh laryngoscope were 37 +/- 12s, 33 +/- 9s, and 30 +/- 12s, respectively. There were no differences among the three devices. CCONCLUSIONS: When operated by experienced anesthesiologists, both Pentax-AWS and Glidescope showed similar performance for nasotracheal intubation compared with Macintosh laryngoscope in normal airway patients.

Obesity-induced asthma: Role of free fatty acid receptors.

Obesity is a major risk factor for the development of asthma, and worsens the key features of asthma including airway hyperresponsiveness, inflammation, and airway remodeling. Although pro- and anti-inflammatory adipocytokines may contribute to the pathogenesis of asthma in obesity, the mechanistic basis for the relationship between asthma and obesity remains unclear. In obese individuals, the increased amount of adipose tissue results in the release of more long-chain free fatty acids as compared to lean individuals, causing an elevation in plasma long-chain free fatty acid concentrations. Recent findings suggest that the free fatty acid receptor 1 (FFAR1), which is a sensor of medium- and long-chain free fatty acids, is expressed on airway smooth muscle and plays a pivotal role in airway contraction and airway smooth muscle cell proliferation. In contrast, FFAR4, which is a sensor for long-chain n-3 polyunsaturated fatty acids and also expressed on airway smooth muscle, does not contribute to airway contraction and airway smooth muscle cell proliferation. Functional roles for short-chain fatty acid receptors FFAR2 and FFAR3 in the pathogenesis of asthma is still under debate. Taken together, adipose-derived long-chain free fatty acids may contribute to the pathogenesis of asthma in obesity through FFAR1.