Ambroxol-enhanced ciliary beating via voltage-gated Ca2+ channels in mouse airway ciliated cells.

Ambroxol (ABX) facilitates the mucociliary clearance (MC) by enhancing ciliary beating in airways. In this study, we focused on airway ciliary beating enhanced by ABX. However, little is known about the ABX-stimulated Ca2+ signalling activating airway ciliary beating. Airway ciliated cells isolated from mice lungs were observed by a high-speed video microscope, and the activities of beating cilia were assessed by CBF (ciliary beat frequency) and CBD (ciliary bend distance, an index of amplitude). ABX (10 µM) enhanced the CBF and CBD by 30%, and the enhancement was inhibited by nifedipine (20 µM, a L-type voltage-gated Ca2+ channel (CaV) inhibitor), or a Ca2+-free solution (approximately 50%). Pre-treatment with BAPTA-AM (10 µM, a chelator of intracellular Ca2+) abolished ABX-stimulated increases in CBF, CBD and [Ca2+]i. Thus, ABX increases [Ca2+]i (intracellular Ca2+ concentration) by stimulating Ca2+ release from the internal stores and nifedipine-sensitive Ca2+ entry. A previous study demonstrated the expression of CaV1.2 in airway cilia. ABX enhanced CBF, CBD and [Ca2+]i even in a high extracellular K+ concentration (155.5 mM), suggesting that it activates CaV1.2 except by depolarization. These enhancements were inhibited by nifedipine. In conclusion, ABX, which increases [Ca2+]i by stimulating Ca2+ release from internal stores and Ca2+ entry through CaV1.2s, enhanced CBF and CBD in airway ciliated cells. ABX is a novel agonist that modulates CaV1.2 of airway beating cilia to enhance CBF and CBD.

A high-TC heavy rare earth monoxide semiconductor TbO with a more than half-filled 4f orbital.

Terbium monoxide, TbO, with an unusual valence state of Tb2+ ([Xe]4f85d1) has been known to exist as a gas phase. In this study, we report a solid phase rock-salt structured TbO in the form of an epitaxial thin film. This TbO is a narrow gap ferromagnetic semiconductor with a bandgap of ∼0.1 eV and high Curie temperature of 231 K, and exhibited negative magnetoresistance of -14% at 9 T and 2 K. Its magnetization steeply increased with the appearance of a wasp-waisted hysteresis curve significantly below the Curie temperature at about 20 K. These results suggested that both 4f and 5d magnetic sublattices were formed at a single magnetic Tb2+ site in TbO.

Changes in choroidal circulation hemodynamics during the menstrual cycle in young, healthy women.

PURPOSE: The current study aimed to investigate the time course of changes in choroidal circulation hemodynamics and their relationship to systemic circulation dynamics during the normal menstrual cycle in young, healthy women using laser speckle flowgraphy (LSFG). METHODS: This prospective study included 26 eyes from 13 young, healthy women (21.3 ± 4.0 years) with a normal menstrual cycle and 24 eyes from 12 young, healthy men (21.8 ± 4.4 years) as a control group. The macular mean blur rate (MBR), a quantitative index of relative blood flow velocity in the choroid, was measured using LSFG. MBR, intraocular pressure (IOP), systolic blood pressure (SBP), diastolic blood pressure (DBP), mean blood pressure (MBP), and ocular perfusion pressure (OPP) were evaluated in the late follicular phase and mid-luteal phase in women and at baseline and 10 days after baseline in men, respectively. RESULTS: In the female group, IOP, SBP, DBP, MBP, and OPP values were significantly higher in the mid-luteal phase than those observed in the late follicular phase (P = 0.035, P < 0.001, P = 0.041, P = 0.001, P = 0.014, respectively). The average macular MBR values in the late follicular phase and mid-luteal phase were 12.7 ± 5.3 and 13.7 ± 6.6 (+7.7 ± 19.4%), representing a significant increase in the mid-luteal phase (P = 0.041). The rate of change in MBR exhibited a significant positive correlation with changes in DBP and MBP (R = 0.456, P = 0.019 and R = 0.474, P = 0.014). However, there were no significant changes in any of the factors in the male group during the study period. CONCLUSIONS: Our results suggest that in young, healthy women with a normal menstrual cycle, choroidal blood flow velocity decreases during the late follicular phase and increases during the mid-luteal phase, depending on systemic circulatory dynamics.

Ezrin knockdown reduces procaterol-stimulated ciliary beating without morphological changes in mouse airway cilia.

Mucociliary clearance, which is conducted by beating cilia cooperating with the surface mucous layer, is a major host defense mechanism of the airway epithelium. Ezrin, a crosslinker between membrane proteins and the actin cytoskeleton, is located in microvilli and around the basal bodies in airway ciliary cells. It is also likely that ezrin plays an important role in apical localization of ß2 adrenergic receptor (ß2AR) in airway ciliary cells. Here, we studied the physiological roles of ezrin by using trachea and airway epithelial cells prepared from ezrin-knockdown (Vil2kd/kd) mice. The trachea and airway ciliary cells of Vil2kd/kd mice presented a normal morphology and basal body orientation, suggesting that ezrin is not directly involved in development and planar cell polarity of cilia. Procaterol stimulates ciliary beating (frequency and amplitude) via ß2AR in the airway ciliary cells. In the Vil2kd/kd mice, airway ciliary beating stimulated with procaterol was partly inhibited due to the impairment of cell surface expression of ß2AR. These results suggest that ezrin regulates the beating of airway ciliary cells by promoting the apical surface localization of ß2AR. This article has an associated First Person interview with the first author of the paper.

Stable Isotope Labeling by Carbon-13 in Bacteria Culture for the Analysis of Residual Avermectin Using Stable Isotope Dilution Liquid Chromatography Tandem Mass Spectrometry.

This study describes the development of a new stable isotope labeling method by carbon-13 in bacteria culture (SILCB) for the analysis of residual antibiotics via liquid chromatography with tandem mass spectrometry (LC-MS/MS). Stable isotope dilution (SID) LC-MS/MS with QuEChERS was employed to determine avermectin, particularly avermectin B1a (AV-a) and B1b (AV-b), based on completely 13C-labeled internal standards (13C-ISs) obtained from the SILCB. Our SILCB was developed from an optimal inorganic medium using 13C6-glucose for Streptomyces avermitilis (14893 strain). A rough extract containing 13C-ISs was purified via high-speed countercurrent chromatography with a volatile two-phase solvent system composed of n-hexane/ethyl acetate/methanol/0.5% formic acid in water (7/3/5/5/, V/V). The purified 13C-ISs were evaluated to confirm the presence of completely 13C-labeled ions with m/z 938 > 326 and m/z 923 > 309 for AV-a and AV-b, respectively. The QuEChERS approach with the 13C-ISs procedure achieved acceptable recovery rates in beef meat samples of 99.5 - 100.0% (RSD < 2.0%, n = 6). For the analysis of residual antibiotics in foodstuffs by SID-LC-MS/MS and QuEChERS, the SILCB represents a significant improvement over previous methods suffering from cumbersome sample preparation and matrix effects.

Moesin is involved in microglial activation accompanying morphological changes and reorganization of the actin cytoskeleton.

Moesin is a member of the ezrin, radixin and moesin (ERM) proteins that are involved in the formation and/or maintenance of cortical actin organization through their cross-linking activity between actin filaments and proteins located on the plasma membranes as well as through regulation of small GTPase activities. Microglia, immune cells in the central nervous system, show dynamic reorganization of the actin cytoskeleton in their process elongation and retraction as well as phagocytosis and migration. In microglia, moesin is the predominant ERM protein. Here, we show that microglial activation after systemic lipopolysaccharide application is partly inhibited in moesin knockout (Msn-KO) mice. We prepared primary microglia from wild-type and Msn-KO mice, and studied them to compare their phenotypes accompanying morphological changes and reorganization of the actin cytoskeleton induced by UDP-stimulated phagocytosis and ADP-stimulated migration. The Msn-KO microglia showed higher phagocytotic activity in the absence of UDP, which was not further increased by the treatment with UDP. They also exhibited decreased ADP-stimulated migration activities compared with the wild-type microglia. However, the Msn-KO microglia retained their ability to secrete tumor necrosis factor α and nitric oxide in response to lipopolysaccharide.

Airway Ciliary Beating Affected by the Pcp4 Dose-Dependent [Ca2+]i Increase in Down Syndrome Mice, Ts1Rhr.

In Ts1Rhr, a Down syndrome model mouse, the airway ciliary beatings are impaired; that is, decreases in ciliary beat frequency (CBF) and ciliary bend angle (CBA, an index of ciliary beat amplitude)). A resumption to two copies of the Pcp4 gene on the Ts1Rhr trisomic segment (Ts1Rhr:Pcp4+/+/-) rescues the decreases in CBF and CBA that occur in Ts1Rhr. In airway cilia, upon stimulation with procaterol (a ß2-agonist), the CBF increase is slower over the time course than the CBA increase because of cAMP degradation by Ca2+/calmodulin-dependent phosphodiesterase 1 (PDE1) existing in the metabolon regulating CBF. In Ts1Rhr, procaterol-stimulated CBF increase was much slower over the time course than in the wild-type mouse (Wt) or Ts1Rhr:Pcp4+/+/-. However, in the presence of 8MmIBMX (8-methoxymethyl isobutylmethyl xanthine, an inhibitor of PDE1) or calmidazolium (an inhibitor of calmodulin), in both Wt and Ts1Rhr, procaterol stimulates CBF and CBA increases over a similar time course. Measurements of cAMP revealed that the cAMP contents were lower in Ts1Rhr than in Wt or in Ts1Rhr:Pcp4+/+/-, suggesting the activation of PDE1A that is present in Ts1Rhr airway cilia. Measurements of the intracellular Ca2+ concentration ([Ca2+]i) in airway ciliary cells revealed that temperature (increasing from 25 to 37 °C) or 4αPDD (a selective transient receptor potential vanilloid 4 (TRPV4) agonist) stimulates a larger [Ca2+]i increase in Ts1Rhr than in Wt or Ts1Rhr:Pcp4+/+/-. In airway ciliary cells of Ts1Rhr, Pcp4-dose dependent activation of TRPV4 appears to induce an increase in the basal [Ca2+]i. In early embryonic day mice, a basal [Ca2+]i increased by PCP4 expressed may affect axonemal regulatory complexes regulated by the Ca2+-signal in Ts1Rhr, leading to a decrease in the basal CBF and CBA of airway cilia.

Isolation and characterization of microsatellite markers from the great hornbill, Buceros bicornis.

Thirteen polymorphic microsatellite markers were isolated and characterized from the great hornbill, Buceros bicornis. In analyses of 20 individuals, the numbers of alleles per locus varied from two to 11. The expected and observed heterozygosities ranged from 0.22 to 0.88 and from 0.20 to 1.00, respectively. The mean polymorphic information content was 0.62. Among these, three loci deviated from the Hardy-Weinberg equilibrium. However, no significant genotypic disequilibrium was detected between any pair of loci. These microsatellite markers are useful for the population genetic study of the great hornbill.